New CGRP-antagonists, process for preparing them and their use as pharmaceutical compositions

ABSTRACT

The present invention relates to the CGRP-antagonists of general formula I 
     
       
         
         
             
             
         
       
     
     wherein R 1 , R 2 , R 3 , R 4  and X are defined as in claim  1 , the tautomers, the isomers, the diastereomers, the enantiomers, the hydrates, mixtures and salts thereof and the hydrates of the salts, particularly the physiologically acceptable salts thereof with inorganic or organic acids or bases, as well as those compounds of general formula I in which one or more hydrogen atoms are replaced by deuterium, pharmaceutical compositions containing these compounds, the use thereof and processes for the preparation thereof.

RELATED APPLICATIONS

This application is a division of U.S. application Ser. No. 12/363,175,filed on Jan. 30, 2009, which is a division of U.S. application Ser. No.11/277,177, filed on Mar. 22, 2006, now U.S. Pat. No. 7,528,129, whichissued on May 5, 2009, which claims benefit, as does the presentapplication, to EP05021283, Filed Sep. 29, 2005, PCT/EP05/04104, filedApr. 18, 2005, PCT/EP05/03094, filed Mar. 23, 2005, AR050101139, filedMar. 23, 2005, and PE000336-2005/01N, filed Mar. 23, 2005.

FIELD OF THE INVENTION

The present invention relates to the CGRP-antagonists of general formulaI

wherein R¹, R², R³, R⁴ and X are as defined hereinbelow, the tautomers,the isomers, the diastereomers, the enantiomers, the hydrates, mixturesand salts thereof and the hydrates of the salts, particularly thephysiologically acceptable salts thereof with inorganic or organic acidsor bases, as well as those compounds of general formula I in which oneor more hydrogen atoms are replaced by deuterium, pharmaceuticalcompositions containing these compounds, the use thereof and processesfor the preparation thereof.

PRIOR ART

International Patent Applications PCT/EP97/04862 and PCT/EP03/11762 havealready described CGRP-antagonists for the treatment of migraine.

DETAILED DESCRIPTION OF THE INVENTION

In the above general formula I in a first embodiment

-   X denotes CH₂, NH, C₁₋₃-alkyl-N, O or S,-   R¹ denotes a group selected from

wherein

-   -   R^(1.1) denotes H, halogen, HO, F₃C or C₁₋₆-alkyl-O,

-   R² denotes a group of general formulae II

wherein

-   -   R^(2.1) denotes H, halogen, C₁₋₃-alkyl-O, C₁₋₃-alkyl or F₃C,    -   R^(2.2) denotes H, H₂N, HO, H₃C—O, H—C(O)—O or        C₁₋₃-alkyl-C(O)—O,    -   R^(2.3) denotes H, halogen, C₁₋₃-alkyl or F₃C, or

-   R² denotes a group selected from

wherein

-   -   R^(2.4) denotes H or H₃C,

-   R³ denotes a group of general formulae III

wherein

-   -   R^(3.1) denotes H, C₁₋₃-alkyl or R^(3.1.1)—(O)C,    -   R^(3.1.1) denotes HO or C₁₋₆-alkyl-O,    -   R^(3.2) denotes H or C₁₋₃-alkyl and    -   R^(3.3) denotes a free pair of electrons or the oxygen atom,

-   R⁴ denotes a 4- to 7-membered oxycycloalkyl group optionally    substituted by R^(4.1) and    -   R^(4.1) denotes NC, HO, C₁₋₃-alkyl or C₁₋₃-alkyl-O,        the tautomers, the diastereomers, the enantiomers, the hydrates        thereof, the mixtures thereof and the salts thereof as well as        the hydrates of the salts, particularly the physiologically        acceptable salts thereof with inorganic or organic acids or        bases.

A preferred first embodiment of the present invention comprises thecompounds of the above general formula I, wherein

-   R¹, R², R³ and R⁴ are defined as hereinbefore under the first    embodiment and-   X denotes CH₂, NH or O,    the tautomers, the diastereomers, the enantiomers, the hydrates    thereof, the mixtures thereof and the salts thereof as well as the    hydrates of the salts, particularly the physiologically acceptable    salts thereof with inorganic or organic acids or bases.

A second embodiment of the present invention comprises the compounds ofthe above general formula I, wherein

-   X, R¹, R³ and R⁴ are defined as hereinbefore under the first    embodiment and-   R² denotes a group of general formula II

wherein

-   -   R^(2.1) denotes H, halogen, C₁₋₃-alkyl-O, C₁₋₃-alkyl or F₃C,    -   R^(2.2) denotes H, H₂N, HO, H₃C—O, H—C(O)—O or        C₁₋₃-alkyl-C(O)—O,    -   R^(2.3) denotes H, halogen, C₁₋₃-alkyl or F₃C,        the tautomers, the diastereomers, the enantiomers, the hydrates        thereof, the mixtures thereof and the salts thereof as well as        the hydrates of the salts, particularly the physiologically        acceptable salts thereof with inorganic or organic acids or        bases.

A preferred second embodiment of the present invention comprises thecompounds of the above general formula I, wherein

-   R¹, R², R³ and R⁴ are defined as hereinbefore under the second    embodiment and-   X denotes CH₂, NH or O,    the tautomers, the diastereomers, the enantiomers, the hydrates    thereof, the mixtures thereof and the salts thereof as well as the    hydrates of the salts, particularly the physiologically acceptable    salts thereof with inorganic or organic acids or bases.

A third embodiment of the present invention comprises the compounds ofthe above general formula I, wherein

-   X denotes CH₂, NH or O,-   R¹ denotes a group selected from

wherein

-   -   R^(1.1) denotes H, Cl, Br, HO, F₃C or H₃C—O,

-   R² denotes a group of general formulae II

wherein

-   -   R^(2.1) denotes H, Cl, Br, H₃C—O, H₃C, F₃C or H₃C—H₂C,    -   R^(2.2) denotes H₂N, HO, H₃C—O, H—C(O)—O or H₃C—C(O)—O,    -   R^(2.3) denotes H, Cl, Br, H₃C or F₃C, or

-   R² denotes a group selected from

wherein

-   -   R^(2.4) denotes H or H₃C,

-   R³ denotes a group of general formulae III

wherein

-   -   R^(3.1) denotes H or H₃C,    -   R^(3.2) denotes H or H₃C and    -   R^(3.3) denotes a free pair of electrons or the oxygen atom,

-   R⁴ denotes a 4- to 7-membered oxycycloalkyl group optionally    substituted by R^(4.1) and    -   R^(4.1) denotes HO or C₁₋₃-alkyl,        the tautomers, the diastereomers, the enantiomers, the hydrates        thereof, the mixtures thereof and the salts thereof as well as        the hydrates of the salts, particularly the physiologically        acceptable salts thereof with inorganic or organic acids or        bases.

A preferred third embodiment of the present invention comprises thecompounds of the above general formula I, wherein

-   X, R¹, R² and R³ are defined as hereinbefore under the second    embodiment and-   R⁴ denotes a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

A fourth embodiment of the present invention comprises the compounds ofthe above general formula I, wherein

-   X, R¹, R³ and R⁴ are defined as hereinbefore under the third    embodiment and-   R² denotes a group of general formula II

wherein

-   -   R^(2.1) denotes H, Cl, Br, H₃C—O, H₃C, F₃C or H₃C—H₂C,    -   R^(2.2) denotes H₂N, HO, H₃C—O, H—C(O)—O or H₃C—C(O)—O,    -   R^(2.3) denotes H, Cl, Br, H₃C or F₃C,        the tautomers, the diastereomers, the enantiomers, the hydrates        thereof, the mixtures thereof and the salts thereof as well as        the hydrates of the salts, particularly the physiologically        acceptable salts thereof with inorganic or organic acids or        bases.

A preferred fourth embodiment of the present invention comprises thecompounds of the above general formula I, wherein

-   X, R¹, R² and R³ are defined as hereinbefore under the fourth    embodiment and-   R⁴ denotes a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

A fifth embodiment of the present invention comprises the compounds ofthe above general formula I, wherein

-   X denotes CH₂, NH or O,-   R¹ denotes a group selected from

wherein

-   -   R^(1.1) denotes H or H₃C—O,

-   R² denotes a group selected from

wherein

-   -   R^(2.1) denotes H₃C or F₃C,    -   R^(2.2) denotes H₂N or HO,    -   R^(2.4) denotes H or H₃C,

-   R³ denotes a group selected from

and

-   R⁴ denotes a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

A preferred fifth embodiment of the present invention comprises thecompounds of the above general formula I, wherein

-   R¹, R² and X are defined as hereinbefore under the fifth embodiment    and —R³—R⁴ together denote a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

A sixth embodiment of the present invention comprises the compounds ofthe above general formula I, wherein

-   X, R¹, R³ and R⁴ are defined as hereinbefore under the fifth    embodiment and-   R² denotes a group selected from

wherein

-   -   R^(2.1) denotes H₃C or F₃C,    -   R^(2.2) denotes H₂N or HO,        the tautomers, the diastereomers, the enantiomers, the hydrates        thereof, the mixtures thereof and the salts thereof as well as        the hydrates of the salts, particularly the physiologically        acceptable salts thereof with inorganic or organic acids or        bases.

A preferred sixth embodiment of the present invention comprises thecompounds of the above general formula I, wherein

-   R¹, R² and X are defined as hereinbefore under the sixth embodiment    and —R³—R⁴ together denote a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

A seventh embodiment of the present invention comprises the compounds ofthe above general formula I, wherein

-   R¹ denotes a group selected from

-   R² denotes a group selected from

-   R³ denotes a group selected from

-   R⁴ denotes a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

A preferred seventh embodiment of the present invention comprises thecompounds of the above general formula I, wherein

-   X, R¹, R³ and R⁴ are defined as hereinbefore under the seventh    embodiment and-   R² denotes a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

An eighth embodiment of the present invention comprises the compounds ofthe above general formula I, wherein

-   R¹ denotes a group selected from

-   R² denotes a group selected from

-   R³—R⁴ together denote a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

A preferred eighth embodiment of the present invention comprises thecompounds of the above general formula I, wherein

-   X, R¹, R³ and R⁴ are defined as hereinbefore under the eighth    embodiment and-   R² denotes a group selected from

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

The following compounds may also be mentioned as examples of mostparticularly preferred compounds of the above general formula I:

No. Structure (1)

(2)

(3)

(4)

(5)

(6)

(7)

(8)

(9)

(10)

(11)

(12)

(13)

(14)

(15)

(16)

(17)

(18)

(19)

(20)

(21)

(22)

(23)

(24)

(25)

(26)

(27)

(28)

(29)

(30)

(31)

(32)

(33)

(34)

(35)

(36)

(37)

(38)

(39)

(40)

(41)

(42)

(43)

(44)

(45)

(46)

(47)

(48)

(49)

(50)

(51)

(52)

(53)

(54)

(55)

(56)

(57)

(58)

(59)

(60)

(61)

(62)

(63)

(64)

(65)

(66)

(67)

(68)

(69)

(70)

(71)

(72)

(73)

(74)

(75)

(76)

(77)

(78)

(79)

(80)

(81)

(82)

(83)

(84)

(85)

(84a)

(85a)

(86)

(87)

(88)

(89)

(90)

(91)

(92)

(93)

(94)

(95)

(96)

(97)

(98)

(99)

(100)

(101)

(102)

(103)

(104)

(105)

(106)

(107)

(108)

(109)

(110)

(111)

(112)

(113)

(114)

(115)

(116)

(117)

(118)

(119)

(120)

(121)

(122)

(123)

(124)

(125)

(126)

(127)

(128)

(129)

(130)

(131)

(132)

(133)

(134)

(135)

(136)

(137)

(138)

(139)

(140)

(141)

(142)

(143)

(144)

(145)

(146)

(147)

(148)

(149)

(150)

(151)

(152)

(153)

(154)

(155)

(156)

(157)

(158)

(159)

(160)

(161)

(162)

(163)

(164)

(165)

(166)

(167)

(168)

(169)

(170)

(171)

(172)

(173)

(174)

(175)

(176)

(177)

(178)

(179)

(180)

(181)

(182)

(183)

(184)

(185)

(186)

(187)

(188)

(189)

(190)

(191)

(192)

(193)

(194)

(195)

(196)

(197)

(198)

(199)

(200)

(201)

(202)

(203)

(204)

(205)

(206)

(207)

(208)

(209)

(210)

(211)

(212)

(213)

(214)

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

The following compounds may also be mentioned as more preferredcompounds of the above general formula I:

No. Structure (34)

(88)

(97)

(103)

(112)

(171)

(172)

(173)

(174)

(177)

(178)

(179)

(180)

(181)

(182)

(183)

(184)

(185)

(186)

(187)

(188)

the tautomers, the diastereomers, the enantiomers, the hydrates thereof,the mixtures thereof and the salts thereof as well as the hydrates ofthe salts, particularly the physiologically acceptable salts thereofwith inorganic or organic acids or bases.

TERMS AND DEFINITIONS USED

Unless stated otherwise, all the substituents are independent of oneanother. If, for example, one group has a number of C₁₋₆-alkyl groups assubstituents, in the case of three substituents, independently of oneanother, one C₁₋₃-alkyl may represent methyl, one may represent ethyland one may represent n-propyl or iso-propyl.

Within the scope of this application, in the definition of possiblesubstituents, these may also be represented in the form of a structuralformula. An asterisk (*) in the structural formula of the substituent isto be understood as being the linking point to the rest of the molecule.

Also included in the subject matter of this invention are the compoundsaccording to the invention, including the salts thereof, in which one ormore hydrogen atoms, for example one, two, three, four or five hydrogenatoms, are replaced by deuterium.

By the term “C₁₋₃-alkyl” (including those which are part of othergroups) are meant branched and unbranched alkyl groups with 1 to 3carbon atoms and by the term “C₁₋₆-alkyl” are meant branched andunbranched alkyl groups with 1 to 6 carbon atoms. Examples include:methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl,tert-butyl, n-pentyl, iso-pentyl, neo-pentyl or hexyl. The followingabbreviations may optionally also be used for the above-mentionedgroups: Me, Et, n-Pr, i-Pr, n-Bu, i-Bu, t-Bu, etc. Unless statedotherwise, the definitions propyl, butyl, pentyl and hexyl include allthe possible isomeric forms of the groups in question. Thus, forexample, propyl includes n-propyl and iso-propyl, butyl includesiso-butyl, sec-butyl and tert-butyl etc.

By the term “4- to 7-membered oxycycloalkyl group” are meant cycloalkylgroups with 4 to 7 carbon atoms, while in each case a CH₂ group isreplaced by an oxygen atom. Examples include:

The above-mentioned oxycycloalkyl groups may optionally be substitutedby hydroxy or methyl groups.

“Halogen” within the scope of the present invention denotes fluorine,chlorine, bromine or iodine. Unless stated to the contrary, fluorine,chlorine and bromine are regarded as preferred halogens.

Compounds of general formula I may have acid groups, mainly carboxylgroups, and/or basic groups such as e.g. amino functions. Compounds ofgeneral formula I may therefore be present as internal salts, as saltswith pharmaceutically useable inorganic acids such as for examplehydrobromic acid, phosphoric acid, nitric acid, hydrochloric acid,sulphuric acid, methanesulphonic acid, ethanesulphonic acid,benzenesulphonic acid, p-toluenesulphonic acid or organic acids such asfor example malic acid, succinic acid, acetic acid, fumaric acid, maleicacid, mandelic acid, lactic acid, tartaric acid, citric acid or as saltswith pharmaceutically useable bases such as alkali or alkaline earthmetal hydroxides, for example sodium hydroxide or potassium hydroxide,or carbonates, ammonia, zinc or ammonium-hydroxides or organic aminessuch as e.g. diethylamine, triethylamine, ethanolamine, diethanolamine,triethanolamine, cyclohexylamine, dicyclohexylamine, inter alia.

The invention relates to the compounds in question, optionally in theform of the individual optical isomers, mixtures of the individualenantiomers or racemates, in the form of the tautomers as well as in theform of the free bases or the corresponding acid addition salts withpharmacologically acceptable acids—such as for example acid additionsalts with hydrohalic acids—for example hydrochloric or hydrobromic acidor organic acids such as for example oxalic, fumaric, diglycolic ormethanesulphonic acid.

The compounds according to the invention may occur as racemates if theyhave only one chiral element, but they may also be obtained as pureenantiomers, i.e. in the (R) or (S) form. Preferred compounds are thosewhich occur as racemates or as the (R) form (for compounds wherein Xdoes not denote the methylene group) or (S) form (for compounds whereinX denotes the methylene group).

However, the application also includes the individual diastereomericpairs of antipodes or the mixtures thereof which are present when thereis more than one chiral element in the compounds of general formula I,as well as the individual optically active enantiomers of which theabove-mentioned racemates are made up.

Preparation Methods

The compounds of general formula I are prepared by methods known inprinciple. Methods of preparing compounds of general formula I whereinR¹, R², R³ and R⁴ are as hereinbefore defined and wherein X denotes the—CH₂, —NH or —C₁₋₃-alkylenyl-N group have already been described inInternational Patent Applications PCT/EP97/04862 and PCT/EP03/11762.Methods of preparing R¹ are also described in International PatentApplications PCT/EP97/04862 and PCT/EP03/11762 as well as in EP 1 619187 A1.

The following methods have proved particularly suitable for preparingthe compounds of general formula I according to the invention wherein Xdenotes the oxygen atom:

(a) In order to prepare compounds of general formula I wherein all thegroups are as hereinbefore defined:coupling a carboxylic acid of general formula IV

wherein R¹ and R² are as hereinbefore defined, with an amine of generalformula V

H—R³—R⁴

wherein R³ and R⁴ are as hereinbefore defined, the link being made viathe nitrogen atom of R³.

Before the reaction is carried out any carboxylic acid functions,primary or secondary amino functions or hydroxy functions present in thegroups of the amine of formula H—R³—R⁴ may be protected by conventionalprotecting groups and any protecting groups used may be cleaved againafter the reaction has been carried out, using methods familiar to theskilled man.

The coupling is preferably carried out using methods known from peptidechemistry (cf. e.g. Houben-Weyl, Methoden der Organischen Chemie, Vol.15/2), for example using carbodiimides such as e.g.dicyclohexylcarbodiimide (DCC), diisopropyl carbodiimide (DIC) orethyl-(3-dimethylaminopropyl)-carbodiimide, O-(1H-benzotriazol-1-yl)N,N,N′,N′-tetramethyluronium hexafluorophosphate (HBTU) ortetrafluoroborate (TBTU) or1H-benzotriazol-1-yl-oxy-tris-(dimethylamino)-phosphoniumhexafluorophosphate (BOP). By adding 1-hydroxybenzotriazole (HOBt) or3-hydroxy-4-oxo-3,4-dihydro-1,2,3-benzotriazine (HOObt) the reactionspeed can be increased. The couplings are normally carried out withequimolar amounts of the coupling components as well as the couplingreagent in solvents such as dichloromethane, tetrahydrofuran,acetonitrile, dimethyl formamide (DMF), dimethyl acetamide (DMA),N-methylpyrrolidone (NMP) or mixtures thereof and at temperaturesbetween −30° C. and +30° C., preferably −20° C. and +25° C. Ifnecessary, N-ethyl-diisopropylamine (Hünig base) is preferably used asan additional auxiliary base.

The so-called “anhydride process” is used as a further coupling methodfor synthesising compounds of general formula I (cf. also: M. Bodanszky,“Peptide Chemistry”, Springer-Verlag 1988, p. 58-59; M. Bodanszky,“Principles of Peptide Synthesis”, Springer-Verlag 1984, p. 21-27). TheVaughan variant of the “mixed anhydride process” is preferred (J. R.Vaughan Jr., J. Amer. Chem. Soc. 73, 3547 (1951)), in which the mixedanhydride of the carboxylic acid of general formula IV which is to becoupled and monoisobutyl carbonate is obtained, using isobutylchlorocarbonate in the presence of bases such as 4-methylmorpholine or4-ethylmorpholine. The preparation of this mixed anhydride and thecoupling with amines of general formula V are carried out in a one-potprocess, using the above-mentioned solvents and at temperatures between−20 and +25° C., preferably between 0° C. and +25° C.

(b) In order to prepare compounds of general formula I wherein all thegroups are as hereinbefore defined:coupling a compound of general formula VI

wherein R¹ and R² are as hereinbefore defined and Nu denotes a leavinggroup, for example a halogen atom, such as the chlorine, bromine oriodine atom, an alkylsulphonyloxy group with 1 to 10 carbon atoms in thealkyl moiety, a phenylsulphonyloxy or naphthylsulphonyloxy groupoptionally mono-, di- or trisubstituted by chlorine or bromine atoms, bymethyl or nitro groups, wherein the substituents may be identical ordifferent, a 1H-imidazol-1-yl, a 1H-pyrazol-1-yl optionally substitutedby one or two methyl groups in the carbon skeleton, a1H-1,2,4-triazol-1-yl, 1H-1,2,3-triazol-1-yl, 1H-1,2,3,4-tetrazol-1-yl,a vinyl, propargyl, p-nitrophenyl, 2,4-dinitrophenyl, trichlorophenyl,pentachlorophenyl, pentafluorophenyl, pyranyl or pyridinyl, adimethylaminyloxy, 2(1H)-oxopyridin-1-yl-oxy,2,5-dioxopyrrolidin-1-yloxy, phthalimidyloxy, 1H-benzotriazol-1-yloxy orazide group, with an amine of general formula V

H—R³—R⁴,

wherein all the groups are as hereinbefore defined and the attachment ismade via the nitrogen atom of the amine R³.

Before the reaction any carboxylic acid functions, primary or secondaryamino functions or hydroxy functions present in the groups of the amineof general formula V may be protected by conventional protecting groupsand after the reaction any protecting groups used may be cleaved againusing methods familiar to those skilled in the art.

The reaction is carried out under Schotten-Baumann or Einhornconditions, i.e. the components are reacted in the presence of at leastone equivalent of an auxiliary base at temperatures between −50° C. and+120° C., preferably −10° C. and +30° C., and optionally in the presenceof solvents. The auxiliary bases used are preferably alkali metal andalkaline earth metal hydroxides, e.g. sodium hydroxide, potassiumhydroxide or barium hydroxide, alkali metal carbonates, e.g. sodiumcarbonate, potassium carbonate or caesium carbonate, alkali metalacetates, e.g. sodium or potassium acetate, as well as tertiary amines,e.g. pyridine, 2,4,6-trimethylpyridine, quinoline, triethylamine,N-ethyl-diisopropylamine, N-ethyl-dicyclohexylamine,1,4-diazabicyclo[2,2,2]octane or 1,8-diazabicyclo[5,4,0]undec-7-ene, thesolvents used may be, for example, dichloromethane, tetrahydrofuran,1,4-dioxane, acetonitrile, dimethyl formamide, dimethyl acetamide,N-methyl-pyrrolidone or mixtures thereof; if alkali metal or alkalineearth metal hydroxides, alkali metal carbonates or acetates are used asthe auxiliary bases, water may also be added to the reaction mixture ascosolvent.

The new compounds of general formula I according to the inventioncontain one or more chiral centres. If for example there are two chiralcentres the compounds may occur in the form of two pairs ofdiastereomeric antipodes. The invention covers the individual isomers aswell as the mixtures thereof.

The diastereomers may be separated on the basis of their differentphysico-chemical properties, e.g. by fractional crystallisation fromsuitable solvents, by high pressure liquid or column chromatography,using chiral or preferably non-chiral stationary phases.

Racemates covered by general formula I may be separated for example byHPLC on suitable chiral stationary phases (e.g. Chiral AGP, ChiralpakAD). Racemates which contain a basic or acidic function can also beseparated via the diastereomeric, optically active salts which areproduced on reacting with an optically active acid, for example (+) or(−)-tartaric acid, (+) or (−)-diacetyl tartaric acid, (+) or(−)-monomethyl tartrate or (+) or (−)-camphorsulphonic acid, or anoptically active base, for example with (R)-(+)-1-phenylethylamine,(S)-(−)-1-phenylethylamine or (S)-brucine.

According to a conventional method of separating isomers, the racemateof a compound of general formula I is reacted with one of theabove-mentioned optically active acids or bases in equimolar amounts ina solvent and the resulting crystalline, diastereomeric, opticallyactive salts thereof are separated using their different solubilities.This reaction may be carried out in any type of solvent provided that itis sufficiently different in terms of the solubility of the salts.Preferably, methanol, ethanol or mixtures thereof, for example in aratio by volume of 50:50, are used. Then each of the optically activesalts is dissolved in water, carefully neutralised with a base such assodium carbonate or potassium carbonate, or with a suitable acid, e.g.dilute hydrochloric acid or aqueous methanesulphonic acid, and in thisway the corresponding free compound is obtained in the (+) or (−) form.

The (R) or (S) enantiomer alone or a mixture of two optically activediastereomeric compounds covered by general formula I may also beobtained by performing the syntheses described above with a suitablereaction component in the (R) or (S) configuration.

The carboxylic acids of general formula IV needed as starting compoundsare obtained by reacting piperidines of general formula VII

wherein R¹ is as hereinbefore defined, with carbonic acid derivatives ofgeneral formula VIII

wherein Y¹ and Y² represent nucleofugic groups, which may be identicalor different, preferably the chlorine atom, the p-nitrophenoxy ortrichloromethoxy group, and with compounds of general formula IX

wherein R² is as hereinbefore defined and Z¹ denotes a protective groupfor a carboxy group, for example a C₁₋₆-alkyl or an optionallysubstituted benzyl group, wherein the alkyl groups may bestraight-chained or branched and the benzyl group may be substituted byone or two methoxy groups.

Preferably Z¹ denotes the methyl, ethyl, tert-butyl or benzyl group.Before the reaction any hydroxy functions present in the group R² of acompound of formula V may be protected by conventional protecting groupsand any protecting groups used may be cleaved again after the reactionby methods familiar to those skilled in the art.

In a first step the compounds of general formula VII are reacted withthe carbonic acid derivatives of general formula VIII in a solvent, forexample in dichloromethane, THF, pyridine or mixtures thereof, at atemperature from −20° C. to 50° C. in the presence of a base, forexample triethylamine, pyridine or ethyldiisopropylamine. The resultingintermediate may be purified or further reacted without purification.

The reaction of these intermediates with compounds of general formula IXis also carried out in one of the above-mentioned solvents, and at thetemperatures specified above, in the presence of a base, such astriethylamine or pyridine, with or without the addition of an activatingreagent such as e.g. 4-dimethylaminopyridine. To activate them thecompounds of general formula IX may also be deprotonated using a metalhydride, such as e.g. NaH or KH, and in this case it is possible todispense with the base or the activating reagent.

The starting compounds of formula VII and VIII are either commerciallyobtainable, known from the literature or may be prepared using methodsknown from the literature.

Methods of preparing compounds of general formula VII are described forexample in International Patent Applications PCT/EP97/04862 andPCT/EP03/11762 and in EP 1 619 187 A1.

One method of obtaining compounds of general formula IX comprisesreacting aldehydes of general formula X

wherein R² is as hereinbefore defined, with N-acetylglycine in aceticanhydride as solvent in the presence of alkali metal acetate, preferablysodium or potassium acetate, at a suitable temperature, preferably at80-130° C.

The azlactones obtained as primary product are hydrolysed without beingisolated to form the compounds of general formula XI

wherein R² is as hereinbefore defined.

Alternatively enamides of the general structure XII

wherein R² is as hereinbefore defined, may be obtained by a couplingreaction of compounds of general formula XIII

Hal-R²,

wherein R² is as hereinbefore defined and Hal denotes the bromine oriodine atom, and methyl 2-acetylaminoacrylate.

The reaction takes place in a suitable solvent such as tetrahydrofuran,dimethylformamide, 1,4-dioxane or acetonitrile, preferably acetonitrile,at temperatures between ambient temperature and 120° C., preferablybetween 50° C. and 80° C., in the presence of a suitable auxiliary basesuch as triethylamine or ethyldiisopropylamine, preferablytriethylamine, and a suitable catalyst system. Suitable catalyst systemsare a combination of a palladium species, such as palladium(II) acetateor bis(acetonitrile)-palladium-dichloride, preferably palladium(II)acetate, and a suitable phosphane ligand, such as triphenyl- ortris-o-tolyl-phosphane, preferably tris-o-tolyl-phosphane.

By further reaction of compounds of general formulae XI and XII in thepresence of aqueous inorganic acids, such as for example sulphuric,phosphoric or hydrochloric acid, but preferably hydrochloric acid,compounds of general formula XIV

are obtained wherein R² is as hereinbefore defined.

These are then converted with suitable reducing agents into thecompounds of general formula XV

wherein R² is as hereinbefore defined.

Suitable reducing agents are alkali metal borohydrides, such as sodiumor potassium borohydride. Other suitable reducing agents arechlorodialkylboranes, such as chlorodicyclohexylborane. If chiralchlorodialkylboranes, such as e.g. B-chlorodiisopinocampheylborane, areused, the compounds of general formula XIII may be isolated inenantiomerically pure form. The further reaction of compounds of generalformula XIII to compounds of general formula IX takes place in analcoholic medium, preferably in methanol or ethanol, in the presence ofa suitable acid, such as hydrochloric acid. The reaction mayalternatively be carried out by reaction in alcoholic solvents,preferably methanol, with thionyl chloride.

All the compounds of general formula I which contain primary orsecondary amino, hydroxy or hydroxycarbonyl functions are preferablyobtained from precursors provided with protective groups. Examples ofprotective groups for amino functions include for example abenzyloxycarbonyl, 2-nitrobenzyloxycarbonyl, 4-nitro-benzyloxycarbonyl,4-methoxy-benzyloxycarbonyl, 2-chloro-benzyloxycarbonyl,3-chloro-benzyloxycarbonyl, 4-chloro-benzyloxycarbonyl,4-biphenylyl-α,α-dimethyl-benzyloxycarbonyl or3,5-dimethoxy-α,α-dimethyl-benzyloxycarbonyl group, an alkoxycarbonylgroup with a total of 1 to 5 carbon atoms in the alkyl moiety, forexample the methoxycarbonyl, ethoxycarbonyl, n-propoxycarbonyl,isopropoxycarbonyl, n-butoxycarbonyl, 1-methyl propoxycarbonyl,2-methylpropoxy-carbonyl or tert-butyloxycarbonyl group, theallyloxycarbonyl, 2,2,2-trichloro-(1,1-dimethylethoxy)carbonyl or9-fluorenylmethoxycarbonyl group or a formyl, acetyl or trifluoroacetylgroup.

Examples of protective groups for hydroxy functions include atrimethylsilyl, triethylsilyl, triisopropyl, tert-butyldimethylsilyl ortert-butyldiphenylsilyl group, a tert-butyl, benzyl, 4-methoxybenzyl or3,4-dimethoxybenzyl group.

A protective group for hydroxycarbonyl functions might be, for example,an alkyl group with a total of 1 to 5 carbon atoms, for example themethyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, allyl,2,2,2-trichloroethyl, benzyl or 4-methoxybenzyl group.

The new compounds of general formula I and the physiologicallyacceptable salts thereof have valuable pharmacological properties, basedon their selective CGRP-antagonistic properties. The invention furtherrelates to pharmaceutical compositions containing these compounds, theiruse and the preparation thereof.

The new compounds mentioned above and the physiologically acceptablesalts thereof have CGRP-antagonistic properties and exhibit goodaffinities in CGRP receptor binding studies. The compounds displayCGRP-antagonistic properties in the pharmacological test systemsdescribed hereinafter.

The following experiments were carried out to demonstrate the affinityof the above-mentioned compounds for human CGRP-receptors and theirantagonistic properties:

A. Binding Studies with SK-N-MC Cells (Expressing the Human CGRPReceptor)

SK-N-MC cells are cultivated in “Dulbecco's modified Eagle medium”. Themedium is removed from confluent cultures. The cells are washed twicewith PBS buffer (Gibco 041-04190 M), detached by the addition of PBSbuffer mixed with 0.02% EDTA, and isolated by centrifuging. Afterresuspension in 20 ml of “Balanced Salts Solution” [BSS (in mM): NaCl120, KCl 5.4, NaHCO₃ 16.2, MgSO₄ 0.8, NaHPO₄ 1.0, CaCl₂ 1.8, D-glucose5.5, HEPES 30, pH 7.40] the cells are centrifuged twice at 100×g andresuspended in BSS. After the number of cells has been determined, thecells are homogenised using an Ultra-Turrax and centrifuged for 10minutes at 3000×g. The supernatant is discarded and the pellet isrecentrifuged in Tris buffer (10 mM Tris, 50 mM NaCl, 5 mM MgCl₂, 1 mMEDTA, pH 7.40) enriched with 1% bovine serum albumin and 0.1%bacitracin, and resuspended (1 ml/1000000 cells). The homogenisedproduct is frozen at −80° C. The membrane preparations are stable formore than 6 weeks under these conditions.

After thawing, the homogenised product is diluted 1:10 with assay buffer(50 mM Tris, 150 mM NaCl, 5 mM MgCl₂, 1 mM EDTA, pH 7.40) andhomogenised for 30 seconds with an Ultra-Turrax. 230 μl of thehomogenised product are incubated for 180 minutes at ambient temperaturewith 50 pM ¹²⁵I-iodotyrosyl-Calcitonin-Gene-Related Peptide (Amersham)and increasing concentrations of the test substances in a total volumeof 250 μl. The incubation is ended by rapid filtration through GF/Bglass fibre filters treated with polyethyleneimine (0.1%) using a cellharvester. The protein-bound radioactivity is measured using a gammacounter. Non-specific binding is defined as the bound radioactivity inthe presence of 1 μM human CGRP-alpha during incubation.

The concentration binding curves are analysed using computer-aidednon-linear curve matching.

The compounds mentioned hereinbefore show IC₅₀ values≦10000 nM in thetest described.

B. CGRP Antagonism in SK-N-MC Cells

SK-N-MC cells (1 million cells) are washed twice with 250 μl incubationbuffer (Hanks' HEPES, 1 mM 3-isobutyl-1-methylxanthine, 1% BSA, pH 7.4)and pre-incubated at 37° C. for 15 minutes. After the addition of CGRP(10 μl) as agonist in increasing concentrations (10⁻¹¹ to 10⁻⁶ M), oradditionally the substance in 3 to 4 different concentrations, themixture is incubated for another 15 minutes.

Intracellular cAMP is then extracted by the addition of 20 μl of 1M HCland centrifugation (2000×g, 4° C., for 15 minutes). The supernatants arefrozen in liquid nitrogen and stored at −20° C.

The cAMP contents of the samples are determined by radioimmunoassay(Messrs. Amersham) and the pA₂ values of antagonistically actingsubstances are determined graphically.

The compounds according to the invention exhibit CGRP-antagonisticproperties in the in vitro test model described, in a dosage rangebetween 10⁻¹² and 10⁻⁶ M.

Types of Indications

In view of their pharmacological properties the compounds according tothe invention and the salts thereof with physiologically acceptableacids are thus suitable for the acute and prophylactic treatment ofheadaches, particularly migraine, cluster headaches and tensionheadaches. Moreover, the compounds according to the invention also havea positive effect on the following diseases: non-insulin-dependentdiabetes mellitus (“NIDDM”), cardiovascular diseases, morphinetolerance, diarrhoea caused by clostridium toxin, skin diseases,particularly thermal and radiation-induced skin damage includingsunburn, lichen, prurigo, pruriginous toxidermies and severe itching,inflammatory diseases, e.g. inflammatory diseases of the joints(osteoarthritis, rheumatoid arthritis, neurogenic arthritis),generalised soft tissue rheumatism (fibromyalgia), neurogenicinflammation of the oral mucosa, inflammatory lung diseases, allergicrhinitis, asthma, COPD, diseases accompanied by excessive vasodilatationand resultant reduced blood supply to the tissues, e.g. shock andsepsis, chronic pain, such as e.g. diabetic neuropathies, neuropathiesinduced by chemotherapy, HIV-induced neuropathies, post-herpeticneuropathies, neuropathies induced by tissue trauma, trigeminalneuralgias, temporomandibular dysfunctions, CRPS (complex regional painsyndrome), back pain and visceral diseases such as for example irritablebowel syndrome (IBS), inflammatory bowel syndrome. In addition, thecompounds according to the invention have a general pain-relievingeffect. The symptoms of menopausal hot flushes caused by vasodilatationand increased blood flow in oestrogen-deficient women andhormone-treated patients with prostate carcinoma and castrated men arefavourably affected by the CGRP antagonists of the present applicationin a preventive and acute-therapeutic capacity, this therapeuticapproach being distinguished from hormone replacement by the absence ofside effects.

Preferably, the compounds according to the invention are suitable forthe acute and preventive treatment of migraine and cluster headaches,for treating irritable bowel syndrome (IBS) and for the preventive andacute therapeutic treatment of hot flushes in oestrogen-deficient women.

The dosage required to achieve a corresponding effect is conveniently0.0001 to 3 mg/kg of body weight, preferably 0.01 to 1 mg/kg of bodyweight, when administered intravenously or subcutaneously, and 0.01 to10 mg/kg of body weight, preferably 0.1 to 10 mg/kg of body weight whenadministered orally, nasally or by inhalation, 1 to 3× a day in eachcase.

If the treatment with CGRP antagonists and/or CGRP release inhibitors isgiven as a supplement to conventional hormone replacement, it isadvisable to reduce the doses specified above, in which case the dosagemay be from 1/5 of the lower limits mentioned above up to 1/1 of theupper limits specified.

The invention further relates to the use of the compounds according tothe invention as valuable adjuvants for the production and purification(by affinity chromatography) of antibodies as well as in RIA and ELISAassays, after suitable radioactive labelling, for example by tritiationof suitable precursors, for example by catalytic hydrogenation withtritium or replacing halogen atoms with tritium, and as a diagnostic oranalytical adjuvant in neurotransmitter research.

Combinations

Categories of active substance which may be used in the combinationinclude e.g. antiemetics, prokinetics, neuroleptics, antidepressants,neurokinin antagonists, anticonvulsants, histamine-H1-receptorantagonists, β-blockers, α-agonists and α-antagonists, ergot alkaloids,mild analgesics, non-steroidal antiinflammatories, corticosteroids,calcium antagonists, 5-HT_(1B/1D)-agonists or other anti-migraine agentswhich may be formulated together with one or more inert conventionalcarriers and/or diluents, e.g. with corn starch, lactose, glucose,microcrystalline cellulose, magnesium stearate, polyvinyl pyrrolidone,citric acid, tartaric acid, water, water/ethanol, water/glycerol,water/sorbitol, water/polyethylene glycol, propylene glycol,cetylstearyl alcohol, carboxymethylcellulose or fatty substances such ashard fat or suitable mixtures thereof, into conventional galenicpreparations such as plain or coated tablets, capsules, powders,suspensions, solutions, metered dose aerosols or suppositories.

Thus other active substances which may be used for the combinationsmentioned above include for example the non-steroidal antiinflammatoriesaceclofenac, acemetacin, acetylsalicylic acid, acetaminophen(paracetamol), azathioprine, diclofenac, diflunisal, fenbufen,fenoprofen, flurbiprofen, ibuprofen, indometacin, ketoprofen,leflunomide, lornoxicam, mefenamic acid, naproxen, phenylbutazone,piroxicam, sulphasalazine, zomepirac or the pharmaceutically acceptablesalts thereof as well as meloxicam and other selective COX₂-inhibitors,such as for example rofecoxib, valdecoxib, parecoxib, etoricoxib andcelecoxib, as well as substances which inhibit earlier or later stagesin prostaglandin synthesis or prostaglandin receptor antagonists such ase.g. EP2-receptor antagonists and IP-receptor antagonists.

It is also possible to use ergotamine, dihydroergotamine,metoclopramide, domperidone, diphenhydramine, cyclizine, promethazine,chlorpromazine, vigabatrin, timolol, isometheptene, pizotifen, botox,gabapentin, pregabaline, duloxetine, topiramate, riboflavin,montelukast, lisinopril, micardis, prochloroperazine, dexamethasone,flunarizine, dextropropoxyphene, meperidine, metoprolol, propranolol,nadolol, atenolol, clonidine, indoramin, carbamazepine, phenyloin,valproate, amitryptiline, imipramine, venlafaxine, lidocaine ordiltiazem and other 5-HT_(1B/1D)-agonists such as, for example,almotriptan, avitriptan, eletriptan, frovatriptan, naratriptan,rizatriptan, sumatriptan and zolmitriptan.

In addition, CGRP-antagonists may be combined with vanilloid receptorantagonists, such as e.g. VR-1 antagonists, glutamate receptorantagonists, such as e.g. mGlu5-receptor antagonists, mGlu1-receptorantagonists, iGlu5-receptor antagonists, AMPA-receptor antagonists,purine receptor blockers, such as e.g. P2X3 antagonists, NO-synthaseinhibitors, such as e.g. iNOS inhibitors, calcium channel blockers, suchas e.g. PQ-type blockers, N-type blockers, potassium channel openers,such as e.g. KCNQ channel openers, sodium channel blockers, such as e.g.PN3 channel blockers, NMDA-receptor antagonists, acid-sensing ionchannel antagonists, such as e.g. ASIC3 antagonists, bradykinin receptorantagonists such as e.g. B1-receptor antagonists, cannabinoid receptoragonists, such as e.g. CB2 agonists, CB1 agonists, somatostatin receptoragonists, such as e.g. sst2 receptor agonists.

The dosage for these active substances is conveniently 1/5 of thenormally recommended dose to 1/1 of the normally recommended dose, i.e.for example 20 to 100 mg sumatriptan.

Formulations

The compounds prepared according to the invention may be administeredeither on their own or optionally in combination with other activesubstances for the treatment of migraine by intravenous, subcutaneous,intramuscular, intra-articular, intrarectal or intranasal route, byinhalation, topically, transdermally or orally, while aerosolformulations are particularly suitable for inhalation. The combinationsmay be administered either simultaneously or sequentially.

Suitable forms for administration include for example tablets, capsules,solutions, syrups, emulsions or inhalable powders or aerosols. Theproportion of pharmaceutically active compound or compounds should be inthe range from 0.1 to 90% by weight, preferably 0.5 to 50% by weight ofthe total composition, i.e. in amounts which are sufficient to achievethe dosage range mentioned hereinbefore.

The preparations may be given orally in the form of tablets, powders,powders in capsules (e.g. hard gelatine capsules), or as solutions orsuspensions. When taken by inhalation the active substance combinationmay be administered as a powder, an aqueous or aqueous-ethanolicsolution or by means of a propellant gas formulation.

Preferably, therefore, pharmaceutical formulations are characterised inthat they contain one or more compounds of formula I according to thepreferred embodiments described hereinbefore.

It is particularly preferable if the compounds of formula I areadministered orally, and it is most preferable if they are administeredonce or twice a day. Suitable tablets may be obtained, for example, bymixing the active substance(s) with known excipients, for example inertdiluents such as calcium carbonate, calcium phosphate or lactose,disintegrants such as corn starch or alginic acid, binders such asstarch or gelatine, lubricants such as magnesium stearate or talc and/oragents for delaying release, such as carboxymethyl cellulose, celluloseacetate phthalate, or polyvinyl acetate. The tablets may also compriseseveral layers.

Coated tablets may be prepared accordingly by coating cores producedanalogously to the tablets with substances normally used for tabletcoatings, for example collidone or shellac, gum arabic, talc, titaniumdioxide or sugar. To achieve delayed release or preventincompatibilities the core may also consist of a number of layers.Similarly the tablet coating may consist of a number or layers toachieve delayed release, possibly using the excipients mentioned abovefor the tablets.

Syrups containing the active substances or combinations thereofaccording to the invention may additionally contain a sweetener such assaccharine, cyclamate, glycerol or sugar and a flavour enhancer, e.g. aflavouring such as vanillin or orange extract. They may also containsuspension adjuvants or thickeners such as sodium carboxymethylcellulose, wetting agents such as, for example, condensation products offatty alcohols with ethylene oxide, or preservatives such asp-hydroxybenzoates.

Capsules containing one or more active substances or combinations ofactive substances may for example be prepared by mixing the activesubstances with inert carriers such as lactose or sorbitol and packingthem into gelatine capsules. Suitable suppositories may be made forexample by mixing with carriers provided for this purpose, such asneutral fats or polyethyleneglycol or the derivatives thereof.

Excipients which may be used include, for example, water,pharmaceutically acceptable organic solvents such as paraffins (e.g.petroleum fractions), vegetable oils (e.g. groundnut or sesame oil),mono- or polyfunctional alcohols (e.g. ethanol or glycerol), carrierssuch as e.g. natural mineral powders (e.g. kaolins, clays, talc, chalk),synthetic mineral powders (e.g. highly dispersed silicic acid andsilicates), sugars (e.g. cane sugar, lactose and glucose), emulsifiers(e.g. lignin, spent sulphite liquors, methylcellulose, starch andpolyvinylpyrrolidone) and lubricants (e.g. magnesium stearate, talc,stearic acid and sodium lauryl sulphate).

For oral use the tablets may obviously contain, in addition to thecarriers specified, additives such as sodium citrate, calcium carbonateand dicalcium phosphate together with various additional substances suchas starch, preferably potato starch, gelatin and the like. Lubricantssuch as magnesium stearate, sodium lauryl sulphate and talc may also beused to produce the tablets. In the case of aqueous suspensions theactive substances may be combined with various flavour enhancers orcolourings in addition to the abovementioned excipients.

It is also preferred if the compounds of formula I are administered byinhalation, particularly preferably if they are administered once ortwice a day. For this purpose, the compounds of formula I have to bemade available in forms suitable for inhalation. Inhalable preparationsinclude inhalable powders, propellant-containing metered-dose aerosolsor propellant-free inhalable solutions which are optionally in admixturewith conventional physiologically acceptable excipients.

Within the scope of the present invention, the term propellant-freeinhalable solutions also includes concentrates or sterile inhalablesolutions ready for use. The formulations which may be used within thescope of the present invention are described in more detail in the nextpart of the specification.

Inhalable Powders

If the compounds of formula I are present in admixture withphysiologically acceptable excipients, the following physiologicallyacceptable excipients may be used to prepare the inhalable powdersaccording to the invention: monosaccharides (e.g. glucose or arabinose),disaccharides (e.g. lactose, saccharose, maltose), oligo- andpolysaccharides (e.g. dextrans), polyalcohols (e.g. sorbitol, mannitol,xylitol), salts (e.g. sodium chloride, calcium carbonate) or mixtures ofthese excipients with one another. Preferably, mono- or disaccharidesare used, while the use of lactose or glucose is preferred,particularly, but not exclusively, in the form of their hydrates. Forthe purposes of the invention, lactose is the particularly preferredexcipient, while lactose monohydrate is most particularly preferred.Processes for producing the inhalable powders according to the inventionby grinding and micronising and by finally mixing the ingredientstogether are known from the prior art.

Propellant-Containing Aerosols for Inhalation

The inhalation aerosols containing propellant gas which may be usedaccording to the invention may contain I dissolved in the propellant gasor in dispersed form. The propellant gases which may be used to preparethe inhalation aerosols according to the invention are known from theprior art. Suitable propellant gases are selected from amonghydrocarbons such as n-propane, n-butane or isobutane andhalohydrocarbons such as preferably fluorinated derivatives of methane,ethane, propane, butane, cyclopropane or cyclobutane. The propellantgases mentioned above may be used on their own or in mixtures thereof.Particularly preferred propellant gases are fluorinated alkanederivatives selected from TG134a (1,1,1,2-tetrafluoroethane), TG227(1,1,1,2,3,3,3-heptafluoropropane) and mixtures thereof. Thepropellant-driven inhalation aerosols which may be used for the purposeaccording to the invention may also contain other ingredients such asco-solvents, stabilisers, surfactants, antioxidants, lubricants and pHadjusters. All these ingredients are known in the art.

Propellant-Free Inhalable Solutions

The compounds of formula I are preferably used to preparepropellant-free inhalable solutions and inhalable suspensions. Thesolvent used may be an aqueous or alcoholic, preferably an ethanolicsolution. The solvent may be water on its own or a mixture of water andethanol. The solutions or suspensions are adjusted to a pH of 2 to 7,preferably 2 to 5, using suitable acids. The pH may be adjusted usingacids selected from inorganic or organic acids. Examples of particularlysuitable inorganic acids include hydrochloric acid, hydrobromic acid,nitric acid, sulphuric acid and/or phosphoric acid. Examples ofparticularly suitable organic acids include ascorbic acid, citric acid,malic acid, tartaric acid, maleic acid, succinic acid, fumaric acid,acetic acid, formic acid and/or propionic acid etc. Preferred inorganicacids are hydrochloric and sulphuric acids. It is also possible to usethe acids which have already formed an acid addition salt with one ofthe active substances. Of the organic acids, ascorbic acid, fumaric acidand citric acid are preferred. If desired, mixtures of the above acidsmay be used, particularly in the case of acids which have otherproperties in addition to their acidifying qualities, e.g. asflavourings, antioxidants or complexing agents, such as citric acid orascorbic acid, for example. According to the invention, it isparticularly preferred to use hydrochloric acid to adjust the pH.

Co-solvents and/or other excipients may be added to the propellant-freeinhalable solutions according to the invention. Preferred co-solventsare those which contain hydroxyl groups or other polar groups, e.g.alcohols—particularly isopropyl alcohol, glycols—particularlypropyleneglycol, polyethyleneglycol, polypropyleneglycol, glycolether,glycerol, polyoxyethylene alcohols and polyoxyethylene fatty acidesters.

The terms excipients and additives in this context denote anypharmacologically acceptable substance which is not an active substancebut which can be formulated with the active substance or substances inthe pharmacologically suitable solvent in order to improve thequalitative properties of the active substance formulation. Preferably,these substances have no pharmacological effect or, in connection withthe desired therapy, no appreciable or at least no undesirablepharmacological effect. The excipients and additives include, forexample, surfactants such as soya lecithin, oleic acid, sorbitan esters,such as polysorbates, polyvinylpyrrolidone, other stabilisers,complexing agents, antioxidants and/or preservatives which guarantee orprolong the shelf life of the finished pharmaceutical formulation,flavourings, vitamins and/or other additives known in the art. Theadditives also include pharmacologically unobjectionable salts such assodium chloride as isotonic agents. The preferred excipients includeantioxidants such as ascorbic acid, for example, provided that it hasnot already been used to adjust the pH, vitamin A, vitamin E,tocopherols and similar vitamins and provitamins occurring in the humanbody. Preservatives may be used to protect the formulation fromcontamination with pathogens. Suitable preservatives are those which areknown in the art, particularly cetyl pyridinium chloride, benzalkoniumchloride or benzoic acid or benzoates such as sodium benzoate in theconcentration known from the prior art.

Experimental Section

As a rule, IR, ¹H-NMR and/or mass spectra have been obtained for thecompounds prepared. Unless otherwise stated, R_(f) values are obtainedusing ready-made silica gel TLC plates 60 F254 (E. Merck, Darmstadt,Item no. 1.05714) without chamber saturation. The R_(f) valuesdetermined under the heading Polygram-Alox are obtained using ready-madePolygram Alox N/UV₂₅₄ TLC films (coated with 0.2 mm aluminium oxide)made by Macherey-Nagel (Düren, Item No. 802 021). The ratios given forthe eluants relate to units by volume of the solvents in question. Theunits by volume specified for NH₃ refer to a concentrated solution ofNH₃ in water.

Unless otherwise stated, the acid, base and salt solutions used forworking up the reaction solutions are aqueous systems of the specifiedconcentrations. For chromatographic purification, silica gel made byMillipore (MATREX™, 35-70 μm) is used. For chromatographic purification,aluminium oxide (Alox) made by ICN Biomedicals (Eschwege, Item number02090) is used. According to the manufacturer's instructions therequired activity stage is produced before use. The HPLC data given aremeasured using the parameters shown below:

Method A:

percent by percent by volume of volume of water acetonitrile time (with0.1% (with 0.1% (min) formic acid) formic acid) 0 95 5 4.5 10 90 5 10 905.5 90 10Analytical column: Zorbax column (Agilent Technologies), SB (StableBond) C18; 3.5 μm; 4.6×75 mm; column temperature: 30° C.; flow: 1.6mL/min; injection volume: 5 μL; detection at 254 nm

Method B:

percent by percent by volume of volume of water acetonitrile time (with0.1% (with 0.1% (min) formic acid) formic acid) 0 95 5 9 10 90 10 10 9011 95 5Analytical column: Zorbax column (Agilent Technologies), SB (StableBond) C18; 3.5 μm; 4.6×75 mm; column temperature: 30° C.; flow: 0.8mL/min; injection volume: 5 μL; detection at 254 nm

Method C:

percent by percent by volume of volume of water acetonitrile time (with0.1% (with 0.1% (min) formic acid) formic acid) 0 95 5 4 50 50 4.5 10 905 10 90 5.5 90 10Analytical column: Zorbax column (Agilent Technologies), SB (StableBond) C18; 3.5 μm; 4.6×75 mm; column temperature: 30° C.; flow: 1.6mL/min; injection volume: 5 μL; detection at 254 nm

Method D:

percent by volume percent by volume time of water of acetonitrile (min)(with 0.04% TFA) (with 0.04% TFA) 0 80 20 15 20 80 17 20 80Analytical column: Symmetry C8 Waters—4.6×150 mm; 5 micron, flow: 1.3ml/min, column temperature: 25° C., detection at 254 nm.

Method E:

percent by percent by volume of volume of water acetonitrile time (with0.1% (with 0.1% (min) formic acid) formic acid) 0 95 5 8 50 50 9 10 9010 10 90 11 90 10Analytical column: Zorbax column (Agilent Technologies), SB (StableBond) C18; 3.5 μm; 4.6×75 mm; column temperature: 30° C.; flow: 0.8mL/min; injection volume: 5 μL; detection at 254 nm

Method F:

percent by volume of percent by volume of water acetonitrile time (min)(with 0.1% formic acid) (with 0.1% formic acid) 0 95 5 2 10 90 5 10 905.5 90 10Analytical column: Zorbax column (Agilent Technologies), SB (StableBond) C18; 3.5 μm; 4.6×75 mm; column temperature: 30° C.; flow: 1.6mL/min; injection volume: 5 μL; detection at 254 nm

Method G:

percent by volume of percent by volume of water acetonitrile time (min)(with 0.04% TFA) (with 0.04% TFA) 0 80 20 30 20 80Analytical column: Waters Symmetry C8, 5 μm, 4.6×150 mm; columntemperature: 25° C., flow: 1.3 mL/min, injection volume: 5 μL, detectionat 254 nm.

In preparative HPLC purifications as a rule the same gradients are usedas were used to collect the analytical HPLC data.

The products are collected under mass control and the fractionscontaining the product are combined and freeze-dried.

If no detailed information is given as to the configuration, it is notclear whether it is a pure enantiomer or whether partial or evencomplete racemisation has occurred.

The following abbreviations are used in the description of theexperiments:

-   CDT 1,1′-carbonyl-di-(1,2,4-triazole)-   Cyc cyclohexane-   DCM dichloromethane-   DIPE diisopropylether-   DMF N,N-dimethylformamide-   EtOAc ethyl acetate-   EtOH ethanol-   HATU    O-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium-hexafluoro-phosphate-   AcOH acetic acid-   i.vac. in vacuo (under vacuum)-   MCPBA m-chloroperbenzoic acid-   MeOH methanol-   NaOAc sodium acetate-   PE petroleum ether-   RT ambient temperature-   TBME tert-butylmethylether-   TBTU    O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium-tetrafluoroborate-   TFA trifluoroacetic acid-   THF tetrahydrofuran

Amine A1 1-(4-methyl-tetrahydropyran-4-yl)-piperazine

A1a) 4-(4-benzyl-piperazin-1-yl)-tetrahydropyran-4-carbonitrile

6.5 g (99.8 mmol) potassium cyanide were added to a solution of 10.0 g(96.9 mmol) tetrahydropyran-4-one and 17.1 g (97.0 mmol)1-benzyl-piperazine in 25 mL 4 M HCl and 50 mL water while cooling withice and the reaction mixture was stirred for 40 h at RT. The precipitateformed was suction filtered, washed with water and dried.

Yield: 22.0 g (76% of theory)

ESI-MS: (M+H)⁺=286

A1b) 1-benzyl-4-(4-methyl-tetrahydropyran-4-yl)-piperazine

Under a nitrogen atmosphere 6.00 g (21.02 mmol)4-(4-benzyl-piperazin-1-yl)-tetrahydropyran-4-carbonitrile in 200 mL dryTHF were prepared and 30 mL ethylmagnesium chloride solution (90.0 mmol,3 M in diethyl ether) was added dropwise and the reaction solution wasstirred for 3 h at RT. Saturated NH₄Cl solution was added, the mixturewas stirred for 10 min, extracted exhaustively with diethyl ether andthe combined organic phases were dried over Na₂SO₄. After the desiccantand solvent had been eliminated the residue was purified bychromatography (Alox, activity stage II-III, gradient DCM/MeOH 100:1towards 50:1).

Yield: 4.85 g (84% of theory)

ESI-MS: (M+H)⁺=275

A1c) 1-(4-methyl-tetrahydropyran-4-yl)-piperazine

A suspension of 4.84 g (17.6 mmol)1-benzyl-4-(4-methyl-tetrahydropyran-4-yl)-piperazine and 400 mg 10%Pd/C in 100 mL MeOH was hydrogenated at 50° C. and 3447 hPa hydrogenpressure for 4 h. To complete the reaction 1 mL concentrated HCl wasadded and the mixture was hydrogenated for a further 18 h at 50° C. and3447 hPa hydrogen pressure. The catalyst was suction filtered and thefiltrate was evaporated to dryness. The product, which was obtained asthe hydrochloride salt, was further reacted without purification.

Yield: 4.00 g (100% of theory)

ESI-MS: (M+H)⁺=185

Example 1(R)-1-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)piperidine-1-carboxylate

1a) 4-methyl-3H-benzoxazol-2-one

76.0 g (0.45 mol) CU in 1 L DCM were added dropwise at 0° C. to asolution of 50.0 g (0.39 mol) 5-amino-m-cresol and 210 mLethyldiisopropylamine (1.2 mol) in 1 L DCM. After the end of thereaction the reaction mixture was combined with 250 mL water, theorganic phase was separated off and washed twice with 250 mL 1 M KHSO₄solution and 250 mL water and dried over MgSO₄. After the desiccant andsolvent had been eliminated the residue obtained was dissolved in 200 mLEtOAc, refluxed, combined with 100 mL PE, slowly cooled to RT, theprecipitate formed was suction filtered and dried.

Yield: 39.2 g (67% of theory)

ESI-MS: (M+H)⁺=150

R_(f)=0.65 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

1b) 6-bromo-4-methyl-3H-benzoxazol-2-one

35.8 g (199.1 mmol) N-bromosuccinimide were added to a solution of 29.5g (197.8 mmol) 4-methyl-3H-benzoxazol-2-one in 200 mL AcOH and themixture was stirred overnight at RT. The reaction solution was combinedwith 800 mL water, stirred for 15 min at RT, the precipitate was suctionfiltered, washed with water and dried in the vacuum drying cupboard at60° C.

Yield: 43.0 g (95% of theory)

ESI-MS: (M+H)⁺=226/228 (Br)

R_(f)=0.35 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

1c) methyl(Z,E)-2-acetylamino-3-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-acrylate

Under a nitrogen atmosphere 5.4 g (23.9 mmol) Pd(OAc)₂ and 7.5 g (24.0mmol) tri-o-tolyl-phosphane were added to a solution of 38.3 g (168.0mmol) 6-bromo-4-methyl-3H-benzoxazol-2-one and 28.0 g (191.7 mmol)methyl 2-acetylamino-acrylate in 800 mL acetonitrile and 480 mLtriethylamine, the reaction mixture was stirred for 18 h at 80° C. andthen evaporated down i.vac. The residue was combined with 100 mL waterand 50 mL EtOAc and the precipitate was filtered off. The crystals weredissolved by refluxing in MeOH/DCM 1:1, combined with activatedcharcoal, filtered off and the filtrate was evaporated to dryness.

Yield: 31.2 g (64% of theory)

ESI-MS: (M+H)⁺=291

R_(f)=0.38 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

1d) 3-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-2-oxo-propionic acid

160 mL 4 M HCl were added to a solution of 31.2 g (107.5 mmol) methyl(Z,E)-2-acetylamino-3-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-acrylatein 320 mL 1,4-dioxane and the reaction solution was refluxed for 5 h.The mixture was evaporated down i.vac., the precipitate was filteredoff, washed with water and dried at 60° C. in the vacuum dryingcupboard.

Yield: 24.9 g (98% of theory)

ESI-MS: (M+H)⁺=236

R_(f)=0.38 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

1e)(R)-2-hydroxy-3-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-propionicacid

Under a nitrogen atmosphere a solution of 60.0 g (187.1 mmol)(1R)-B-chloro-diisopinocampheylborane in 200 mL THF was added dropwiseto a solution of 24.9 g (105.9 mmol)3-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-2-oxo-propionic acid and20.0 mL (143.9 mmol) triethylamine in 400 mL THF, cooled to −35° C.,within 15 min and the reaction solution was stirred overnight at RT.Then the reaction solution was carefully made alkaline with 1 M NaOH at5° C., combined with 400 mL EtOAc and stirred for 15 min. The organicphase was separated off, extracted twice with 100 mL of 1 M NaOH andwith 100 mL water. The combined aqueous phases were acidified withsemiconc. HCl and extracted twice with 150 mL EtOAc in each case. Thecombined organic phases were dried over MgSO₄ and evaporated down i.vac.

Yield: 20.8 g (83% of theory)

ESI-MS: (M+H)⁺=238

R_(f)=0.10 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

1f) methyl(R)-2-hydroxy-3-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-propionate

23.0 g (97.0 mmol)(R)-2-hydroxy-3-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-propionicacid were dissolved in 200 mL methanolic HCl (1.3 M), stirred overnightat RT and then evaporated down i.vac. The residue was combined with 200mL EtOAc, washed with 15% K₂CO₃ solution and the organic phase was driedover Na₂SO₄. After the desiccant and solvent had been eliminated theresidue was combined with DIPE, the crystals were filtered off and driedat 50° C. in the vacuum drying cupboard.

Yield: 14.6 g (60% of theory)

ESI-MS: (M+H)⁺=252

R_(f)=0.44 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

1g) (R)-1-carboxy-2-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-ethyl4-(2-oxo-1,2,4,5,-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere 4.1 g (20.1 mmol) 4-nitrophenylchloroformate in 20 mL THF were metered into 40 mL pyridine at a bathtemperature of 60° C. within 10 min, the mixture was stirred for 5 min,then 5.0 g (19.9 mmol) methyl(R)-2-hydroxy-3-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-propionateand 20 mL pyridine were added and the reaction mixture was stirred for1.5 h at 60° C. The reaction solution was combined with 4.9 g (20.0mmol) 3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one andstirred for 2 h at 100° C. After the end of the reaction the mixture wascombined with 150 mL EtOAc, washed three times with 70 mL 1 M KHSO₄solution and 12 times with 50 mL 15% K₂CO₃ solution and the organicphase was dried over MgSO₄. After the desiccant and solvent had beeneliminated the residue was dissolved in 60 mL THF, combined with 250 mgLiOH in 10 mL water and the reaction mixture was stirred for 3 h at RT.The THF was eliminated i.vac., the aqueous residue was combined with 60mL TBME, insoluble ingredients were filtered off, the organic phase wasseparated off and the aqueous phase was acidified with 1 M HCl. After 1h at RT the precipitate formed was suction filtered, washed with waterand dried at 60° C. in the vacuum drying cupboard.

Yield: 2.5 g (25% of theory)

ESI-MS: (M−H)⁻=507

R_(f)=0.10 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

1 h(R)-1-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

30 mg (0.18 mmol) 1-(tetrahydropyran-4-yl)-piperazine were added at RTto a solution of 70 mg (0.14 mmol)(R)-1-carboxy-2-(4-methyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-ethyl4-(2-oxo-1,2,4,5,-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate,50 mg (1.12 mmol) TBTU and 25 μL (0.18 mmol) triethylamine in 1 mL DMFand the reaction solution was stirred overnight. It was purified by HPLCwithout any further working up; the fractions containing the productwere combined and lyophilised.

Yield: 69 mg (68% of theory)

ESI-MS: (M−H)⁻=659

retention time (HPLC): 2.7 min (method A)

Example 2(R)-1-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

2a) 3,4-dimethyl-3H-benzoxazol-2-one

A solution of 10.0 g (67.0 mmol) 4-methyl-3H-benzoxazol-2-one in 200 mLTHF was combined with 8.0 g (70.6 mmol) potassium-tert.-butoxide,stirred for 30 min at RT, then combined with 7.0 mL (110.3 mmol)iodomethane and stirred overnight at RT. The reaction mixture wascombined with 100 mL EtOAc, washed twice with 50 mL saturated NaClsolution, the organic phase was dried over MgSO₄ dried, filtered andevaporated to dryness i.vac. The residue was combined with PE/EtOAc 2:1,the precipitate was suction filtered and dried at 60° C. in the vacuumdrying cupboard.

Yield: 9.0 g (82% of theory)

ESI-MS: (M+H)⁺=164

R_(f)=0.56 (silica gel, PE/EtOAc 2:1)

2b) 6-bromo-3,4-dimethyl-3H-benzoxazol-2-one

11.0 g (60.0 mmol) N-bromosuccinimide were added to a solution of 9.0 g(55.2 mmol) 3,4-dimethyl-3H-benzoxazol-2-one in 50 mL AcOH and thereaction mixture was stirred overnight at RT. The reaction solution wascombined with 300 mL water, stirred for 15 min at RT, the precipitatewas suction filtered, washed with water and dried at 60° C. in thevacuum drying cupboard.

Yield: 12.7 g (95% of theory)

ESI-MS: (M+H)⁺=242/244 (Br)

R_(f)=0.52 (silica gel, PE/EtOAc 2:1)

2c) methyl(Z,E)-2-acetylamino-3-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-acrylate

Under a nitrogen atmosphere 1.8 g (8.0 mmol) Pd(OAc)₂ and 2.5 g (8.0mmol) tri-o-tolyl-phosphane was added to a solution of 13.2 g (54.5mmol) 6-bromo-3,4-dimethyl-3H-benzoxazol-2-one and 9.0 g (61.6 mmol)methyl 2-acetylamino-acrylate in 250 mL acetonitrile and 160 mLtriethylamine and the reaction mixture was stirred for 18 h at 80° C.The reaction solution was evaporated down i.vac., the residue wascombined with 100 mL water and 50 mL EtOAc and the precipitate wasfiltered off. This was dissolved in MeOH/DCM (1:1), combined withactivated charcoal, filtered off and the filtrate was evaporated todryness.

Yield: 8.7 g (52% of theory)

ESI-MS: (M+H)⁺=305

R_(f)=0.47 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

2d) 3-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-2-oxo-propionicacid

40 mL of 4 M HCl were added to a solution of 8.7 g (28.6 mmol) methyl(Z,E)-2-acetylamino-3-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-acrylatein 80 mL of 1,4-dioxane and the reaction solution was refluxed for 5 hand then left overnight at RT. The mixture was evaporated down i.vac.,the precipitated product was filtered off, this was washed with waterand dried at 60° C. in the vacuum drying cupboard.

Yield: 6.6 g (93% of theory)

ESI-MS: (M+H)⁺=250

R_(f)=0.13 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

2e)(R)-3-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-2-hydroxy-propionicacid

Under a nitrogen atmosphere a solution of 15.0 g (46.8 mmol)(1R)-B-chlorodiisopinocampheylborane in 50 mL THF was added dropwisewithin 15 min to a solution of 6.6 g (26.5 mmol)3-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-2-oxo-propionic acidand 5.0 mL (36.0 mmol) triethylamine in 100 mL THF, cooled to −35° C.,and the reaction solution was stirred overnight at RT. Then at 5° C. themixture was combined with 60 mL 1 M NaOH and 100 mL EtOAc, stirred for15 min, the organic phase was separated off and extracted twice with 30mL 1 M NaOH and 40 mL water. The combined aqueous phases were acidifiedwith semiconc. HCl and extracted twice with 100 mL EtOAc. The combinedorganic phases were dried over MgSO₄ and evaporated down i.vac.

Yield: 3.4 g (51% of theory)

ESI-MS: (M+H)⁺=252

R_(f)=0.13 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

2f) methyl(R)-3-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-2-hydroxy-propionate

3.4 g (13.5 mmol)(R)-3-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-2-hydroxy-propionicacid were dissolved in 40 mL methanolic HCl (1.3 M) and the reactionmixture was stirred overnight at RT. The mixture was evaporated downi.vac., the residue was taken up in 200 mL EtOAc, washed with 15% K₂CO₃solution and the organic phase was dried over Na₂SO₄. After thedesiccant and solvent had been eliminated the residue was furtherreacted without purification.

Yield: 2.5 g (70% of theory)

ESI-MS: (M+H)⁺=266

R_(f)=0.54 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

2g)(R)-1-carboxy-2-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-ethyl4-(2-oxo-1,2,4,5,-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere at 60° C., a solution of 2.0 g (10.0 mmol)4-nitrophenyl chloroformate in 10 mL THF was added within 10 min to 20mL pyridine and stirred for 10 min. Then a solution of 2.5 g (9.4 mmol)methyl(R)-3-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-2-hydroxy-propionatein 10 mL pyridine was added, the mixture was stirred for a further 2.5 hat 60° C. and then combined with 2.5 g (10.0 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one. Thereaction solution was stirred for 3 h at 100° C. After the reaction hadended the reaction mixture was evaporated down i.vac., combined 150 mLEtOAc, the organic phase was washed three times with 40 mL 1 M KHSO₄solution and 12 times with 30 mL 15% K₂CO₃ solution and dried overMgSO₄. After the desiccant and solvent had been eliminated the residuewas dissolved in 60 mL THF, combined with 250 mg LiOH in 10 mL water andthe reaction mixture was stirred for 3 h at RT. The THF was eliminatedi.vac., the aqueous phase was diluted with 60 mL EtOAc, filtered toremove any insoluble constituents and the organic phase was separatedoff. The aqueous phase was acidified with 15 mL 1 M HCl, extracted threetimes with 50 mL EtOAc and the combined organic phases were dried overMgSO₄. After the desiccant and solvent had been eliminated the residuewas dissolved in 30 mL isopropanol at 80° C. The solution was left tocool slowly overnight, the precipitate suction filtered, washed withisopropanol and dried at 60° C. in the vacuum drying cupboard.

Yield: 1.1 g (22% of theory)

ESI-MS: (M+H)⁺=523

R_(f)=0.31 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

2h)(R)-1-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 1h from 70 mg (0.13 mmol)(R)-1-carboxy-2-(3,4-dimethyl-2-oxo-2,3-dihydro-benzoxazol-6-yl)-ethyl4-(2-oxo-1,2,4,5,-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 25 mg (0.15 mmol) 1-(tetrahydropyran-4-yl)piperazine.

Yield: 70 mg (66% of theory)

ESI-MS: (M+H)⁺=675

R_(f)=0.63 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

Example 3(R)-1-(7-methyl-1H-benzimidazol-5-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

3a) methyl(Z,E)-2-acetylamino-3-(4-amino-methyl-5-nitro-phenyl)-acrylate

Under an argon atmosphere 0.7 g (2.9 mmol) Pd(OAc)₂ and 0.9 g (2.9 mmol)tri-o-tolyl-phosphane were added to a solution of 9.0 g (39.0 mmol)4-bromo-2-methyl-6-nitro-phenylamine and 10.0 g (69.9 mmol) methyl2-acetylamino-acrylate in 100 mL acetonitrile and 100 mL triethylamine.The reaction mixture was stirred for 24 h at 90° C. bath temperature,evaporated down i.vac., the residue was combined with 200 mL water and200 mL EtOAc and the precipitate was filtered off. The crystals weredissolved in 500 mL MeOH by refluxing, filtered off while hot and thefiltrate was evaporated to dryness i.vac.

Yield: 8.0 g (70% of theory)

ESI-MS: (M+H)⁺=294

3b) 3-(4-amino-3-methyl-5-nitro-phenyl)-2-oxo-propionic acid

60 mL of a 4 M HCl were metered into a solution of 8.0 g (53.1 mmol)methyl (Z,E)-2-acetylamino-3-(4-amino-methyl-5-nitro-phenyl)-acrylate in60 mL 1,4-dioxane, refluxed for 3 h with stirring, the reaction solutionwas evaporated down

i.vac. and the residue was combined with ice. The precipitate wasfiltered off, washed with ice water and dried.

Yield: 6.5 g (95% of theory)

EI-MS: (M)⁺=238

3c) (R)-3-(4-amino-3-methyl-5-nitro-phenyl)-2-hydroxy-propionic acid

Under a nitrogen atmosphere a solution of 12.0 g (37.4 mmol)(1R)-B-chlorodiisopinocampheylborane in 40 mL THF was added dropwisewithin 15 min to a solution of 6.5 g (26.0 mmol)3-(4-amino-3-methyl-5-nitro-phenyl)-2-oxo-propionic acid and 4.5 mL(32.4 mmol) triethylamine in 100 mL THF cooled to −35° C. and thereaction solution was stirred overnight at RT. Then the reactionsolution was carefully combined with 60 mL 1 M NaOH and 150 mL diethylether at 5° C. and stirred for 15 min. The organic phase was separatedoff, extracted three times with 40 mL 1 M NaOH and once with 40 mLwater. The combined aqueous phases were acidified with semiconc. HClwhile cooling with an ice bath and extracted twice with 120 mL EtOAc ineach case. The combined organic phases were dried over Na₂SO₄ andevaporated down i.vac. The crude product was obtained, which was furtherreacted without purification.

Yield: 6.0 g (67% of theory)

3d) methyl (R)-3-(4-amino-3-methyl-5-nitro-phenyl)-2-hydroxy-propionate

4.0 mL (54.8 mmol) SOCl₂ were slowly added dropwise to 90 mL MeOH whilecooling with ice/acetone and at 0° C. 6.0 g (17.5 mmol)(R)-3-(4-amino-3-methyl-5-nitro-phenyl)-2-hydroxy-propionic acid in 10mL MeOH were also added dropwise. The reaction solution was stirred for1 h at 0° C. and for 1 h at RT and then evaporated down i.vac. Theresidue was combined with EtOAc, washed with saturated NaHSO₄ solutionand dried over Na₂SO₄. After the desiccant and solvent had beeneliminated the residue was purified by chromatography (silica gel,gradient DCM/MeOH 100:1 to 50:1).

Yield: 3.4 g (76% of theory)

ESI-MS: (M+H)⁺=255

R_(f)=0.43 (Polygram, DCM/MeOH 50:1)

3e) (R)-2-(4-amino-3-methyl-5-nitro-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere 1.8 g (14.7 mmol) 4-dimethylaminopyridine in25 mL pyridine were first combined with 2.7 g (13.4 mmol) 4-nitrophenylchloroformate while cooling with an ice bath, stirred for 30 min at RT,then combined with 3.4 g (13.2 mmol) methyl(R)-3-(4-amino-3-methyl-5-nitro-phenyl)-2-hydroxy-propionate in 15 mLpyridine, stirred again for 2 h at RT, and then combined with 3.5 g(14.3 mmol) 3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-oneand stirred for 5 h at RT. After the end of the reaction the reactionmixture was evaporated down i.vac., the residue was combined with EtOAc,the organic phase was washed with 10% KHSO₄ solution and saturatedNaHSO₄ solution and dried over Na₂SO₄. After the desiccant and solventhad been eliminated the residue was purified by chromatography (silicagel, DCM/MeOH 25:1).

Yield: 3.7 g (50% of theory)

ESI-MS: (M+H)⁺=526

R_(f)=0.42 (Polygram, DCM/MeOH 25:1)

3f) (R)-1-methoxycarbonyl-2-(7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

1.2 g (2.3 mmol)(R)-2-(4-amino-3-methyl-5-nitro-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatewere dissolved in 50 mL formic acid and combined with 300 mg 10% Pd/C.The mixture was hydrogenated for 2 h in a Parr apparatus at 60° C. undera hydrogen pressure of 3447 hPa. Then the catalyst was filtered off, thefiltrate was evaporated down i.vac. and the residue was purified bychromatography (Alox, gradient DCM/MeOH 40:1 to 30:1).

Yield: 880 mg (76% of theory)

ESI-MS: (M+H)⁺=506

R_(f)=0.40 (Polygram-Alox, DCM/MeOH 25:1)

3g) (R)-1-carboxy-2-(7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 96 mg (4.0 mmol) LiOH in 5 mL water was added dropwise toa solution of 910 mg (1.8 mmol)(R)-1-methoxycarbonyl-2-(7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 12 mL THF and the reaction solution was stirred overnight at RT. Theresidue was combined with 1 mL 4 M HCl and evaporated to dryness i.vac.

Yield: 980 mg (100% of theory)

ESI-MS: (M+H)⁺=492

3h)(R)-1-(7-methyl-1H-benzimidazol-5-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 1h from 120 mg (0.22 mmol)(R)-1-carboxy-2-(7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 50 mg (0.29 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 70 mg (45% of theory)

ESI-MS: (M+H)⁺=644

retention time (HPLC): 2.0 min (method A)

Example 4(R)-1-(2-dimethylamino-7-methyl-1H-benzimidazol-5-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

4a) (R)-2-(3,4-diamino-5-methyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 2.00 g (3.24 mmol)(R)-2-(4-amino-3-methyl-5-nitro-phenyl)-1-methoxy-carbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate(Example 3e, purity 85%) in 100 mL MeOH was hydrogenated for 3.5 h at50° C. and 3447 hPa hydrogen pressure. The catalyst was filtered off andthe filtrate was evaporated down i.vac. The residue was purified bychromatography (Alox, activity stage II-III, gradient DCM/MeOH 30:1 to15:1).

Yield: 1.35 g (84% of theory)

ESI-MS: (M+H)⁺=496

4b)(R)-2-(2-dimethylamino-7-methyl-1H-benzimidazol-5-yl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere 230 mg (0.72 mmol) TBTU was added to asolution of 310 mg (0.63 mmol)(R)-2-(3,4-diamino-5-methyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 140 μL (1.00 mmol) triethylamine in 30 mL 1,4-dioxane and thereaction mixture was refluxed for 12 h. The mixture was evaporated downi.vac., the residue was taken up in EtOAc, the organic phase was washedwith saturated NaHCO₃ solution and dried over Na₂SO₄. After thedesiccant and solvent had been eliminated the residue was purified bychromatography (Alox, activity stage II-III, DCM/MeOH 30:1). Furtherpurification was carried out by silica gel chromatography (gradientDCM/MeOH 12:1 to 6:1).

Yield: 85 mg (25% of theory)

ESI-MS: (M+H)⁺=549

R_(f)=0.50 (Polygram-Alox, DCM/MeOH 25:1)

4c)(R)-1-carboxy-2-(2-dimethylamino-7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 9.0 mg (0.38 mmol) LiOH in 0.5 mL water was added to asolution of 80 mg (0.15 mmol)(R)-2-(2-dimethylamino-7-methyl-1H-benzimidazol-5-yl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 4 mL THF and the reaction mixture was stirred for 4 h at RT. 100 μLof 4 M HCl was added, the mixture was evaporated down i.vac. and thecrude product was dried and then further reacted without purification.

ESI-MS: (M+H)⁺=535

retention time (HPLC): 2.8 min (method A)

4d)(R)-1-(2-dimethylamino-7-methyl-1H-benzimidazol-5-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 1h from 90 mg (0.14 mmol, purity 85%)(R)-1-carboxy-2-(2-dimethylamino-7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 35 mg (0.21 mmol) 1-(tetrahydropyran-4-yl)piperazine.

Yield: 25 mg (25% of theory)

ESI-MS: (M+H)⁺=687

retention time (HPLC): 2.5 min (method A)

Example 5(R)-1-(2-methoxy-7-methyl-1H-benzimidazol-5-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)piperidine-1-carboxylate

5a)(R)-1-methoxycarbonyl-2-(2-methoxy-7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 1.00 g (2.02 mmol)(R)-2-(3,4-diamino-5-methyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate(Example 4a), 2.0 mL (15.0 mmol) tetramethoxymethane and 40 mgp-toluenesulphonic acid-monohydrate in 20 mL MeOH was refluxed for 1 h.The mixture was evaporated down i.vac. and the residue was purified bychromatography (Alox, activity stage II-III, DCM/MeOH 20:1).

Yield: 0.99 g (92% of theory)

ESI-MS: (M+H)⁺=536

5b) (R)-1-carboxy-2-(2-methoxy-7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 100 mg (4.18 mmol) LiOH in 6 mL water was added to asolution of 980 mg (1.83 mmol)(R)-1-methoxycarbonyl-2-(2-methoxy-7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 18 mL THF and the reaction mixture was stirred overnight at RT. 1.05mL of 4 M HCl was added, the THF was evaporated down i.vac., whereby thecrude product was produced in the form of an oil. The water was decantedoff, the residue was dissolved in DCM/MeOH and dried over Na₂SO₄. Afterthe desiccant and solvent had been eliminated the residue was furtherreacted without purification.

Yield: 0.80 g (80% of theory)

ESI-MS: (M−H)⁻=520

5c)(R)-1-(2-methoxy-7-methyl-1H-benzimidazol-5-ylmethyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 1h from 90 mg (0.16 mmol)(R)-1-carboxy-2-(2-methoxy-7-methyl-1H-benzimidazol-5-yl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 40 mg (0.24 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 33 mg (30% of theory)

ESI-MS: (M+H)⁺=674

retention time (HPLC): 2.8 min (method A)

Example 6(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

A solution of 100 mg (0.18 mmol)(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid, 65 mg (0.20 mmol) TBTU and 35 μL (0.18 mmol) triethylamine in 10mL THF was stirred for 1 h at RT. Then 40 mg (0.24 mmol)1-(tetrahydropyran-4-yl)-piperazine was added and the reaction solutionwas stirred overnight. 20 mL semisaturated NaHCO₃ solution was added,extracted twice with 40 mL EtOAc and the combined organic phases weredried over Na₂SO₄. After the desiccant and solvent had been eliminatedthe residue was purified by HPLC; the fractions containing the productwere combined and lyophilised.

Yield: 84 mg (66% of theory)

ESI-MS: (M+H)⁺=705/707 (Cl)

retention time (HPLC): 6.3 min (method B)

Example 7(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

7a) 2-benzyloxy-5-bromo-1,3-dimethylbenzene

39.9 g (286 mmol) K₂CO₃ were added to a solution of 50.0 g (249 mmol)2,6-dimethyl-4-bromophenol in 500 mL DMF and stirred for 20 min. Then34.0 mL (286 mmol) benzyl chloride were slowly added dropwise and thereaction mixture was stirred for 3 h at a bath temperature of 100° C.After the end of the reaction the mixture was poured onto 500 mL waterand exhaustively extracted with EtOAc. The organic phases were combined,dried over Na₂SO₄ and evaporated down i.vac.

Yield: quantitative

GC-MS: (M⁺)=290/292 (Br)

R_(f)=0.87 (silica gel, Cyc/EtOAc 3:1)

7b) methyl 2-acetylamino-3-(4-benzyloxy-3,5-dimethyl-phenyl)-acrylate

Under a nitrogen atmosphere a mixture of 40.0 g (137 mmol)2-benzyloxy-5-bromo-1,3-dimethylbenzene and 24.1 g (165 mmol) methyl2-acetylamino-acrylate in 420 mL triethylamine and 200 mL acetonitrilewas combined with 3.5 g (11.2 mmol) tri-o-tolyl-phosphane and 2.5 g(11.1 mmol) Pd(OAc)₂ and stirred for 18 h at 80° C. The precipitate wassuction filtered, the filtrate was evaporated down i.vac. and combinedwith 800 mL DCM and 800 mL water. The organic phase was separated off,suction filtered through Na₂SO₄, the solvent was eliminated i.vac., theresidue was stirred with EtOAc, suction filtered and dried i.vac.

Yield: 31.1 g (64% of theory)

ESI-MS: (M+H)⁺=354

retention time (HPLC-MS):8.6 min (method B)

7c) 3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-oxo-propionic acid

31.1 g (88.1 mmol) methyl2-acetylamino-3-(4-benzyloxy-3,5-dimethyl-phenyl)-acrylate in 150 mL1,4-dioxane were combined with 125 mL 4 M HCl, refluxed for 7 h andstirred overnight at RT. The precipitate was suction filtered, washedwith water and dried at 45° C. in the vacuum drying cupboard.

Yield: 14.3 g (54% of theory)

EI-MS: (M)⁺=298

retention time (HPLC-MS):9.0 min (method B)

7d) (R)-3-(4-benzoyl-3,5-dimethyl-phenyl)-2-hydroxy-propionic acid

Under a nitrogen atmosphere a solution of 14.3 g (47.8 mmol)3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-oxo-propionic acid and 8.3 mL(59.8 mmol) triethylamine in 170 mL THF at −35° C. was combined with asolution of 22.1 (69.0 mmol) (1R)-B-chlorodiisopinocampheylborane in 70mL THF within 30 min. After the addition had ended the cooling bath wasremoved and the reaction solution was stirred overnight at RT. Thereaction mixture was made alkaline with 70 mL 1 M NaOH at 0° C.,combined with 100 mL TBME, stirred for 15 min and the phases wereseparated. The organic phase was washed with 50 mL water and three timeswith 50 mL 1 M NaOH. The combined aqueous phases were acidified withsemiconc. HCl, exhaustively extracted with EtOAc and the combinedorganic phases were dried over Na₂SO₄. After the desiccant and solventhad been eliminated the residue was further reacted withoutpurification.

Yield: 14.0 g (98% of theory)

ESI-MS: (M−H)⁻=299

retention time (HPLC-MS):7.9 min (method B)

7e) methyl (R)-3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-hydroxy-propionate

2.0 mL (27.4 mmol) SOCl₂ were added dropwise to a solution, cooled to 0°C., of 14.0 g (23.3 mmol)(R)-3-(4-benzoyl-3,5-dimethyl-phenyl)-2-hydroxy-propionic acid in 150 mLMeOH and the reaction mixture was stirred for 1 h at RT. The reactionsolution was evaporated down i.vac. and the residue was purified bychromatography (silica gel, Cyc/EtOAc 3:1).

Yield: 5.7 g (78% of theory)

ESI-MS: (M+NH₄)⁺=332

retention time (HPLC-MS):9.1 min (method B)

7f) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere 1.93 g (9.58 mmol) 4-nitrophenylchloroformate was added to a solution of 1.17 g (9.58 mmol)4-dimethylaminopyridine in 50 mL pyridine, stirred for 1.5 h at RT,combined with 3.0 g (9.58 mmol) methyl(R)-3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-hydroxy-propionate and stirredfor 20 min at RT. Then 2.35 g (9.58 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one were addedand the mixture was stirred for 20 h at RT. The reaction mixture wasevaporated down i.vac., the residue was taken up in EtOAc, the organicphase was washed with 10% KHSO₄ and saturated NaHCO₃ solution and driedover Na₂SO₄. After the desiccant and solvent had been eliminated theresidue was purified by chromatography (silica gel, gradient Cyc/EtOAc1:1 to 1:2).

Yield: 3.21 g (57% of theory)

ESI-MS: (M+H)⁺=586

retention time (HPLC-MS):10.4 min (method B)

7g) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 3.21 g (5.48 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxy-carbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 80 mL THF was combined with a solution of 200 mg (8.35 mmol) LiOH in40 mL water and stirred for 1 h at RT. The reaction mixture wasevaporated down i.vac., the residue was taken up in 100 mL water,acidified with 2 M HCl, the precipitate was suction filtered and driedin the vacuum drying cupboard at 40° C.

Yield: quantitative

ESI-MS: (M+H)⁺=572

retention time (HPLC-MS):9.2 min (method B)

7h) (R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

3.72 g (6.51 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 50 mL DCM were combined with 300 mg 10% Pd/C and shaken at RT and3000 hPa hydrogen until the reaction came to an end. The catalyst wassuction filtered and the solvent evaporated down i.vac. The residue wastriturated with DIPE and suction filtered.

Yield: 2.41 g (77% of theory)

ESI-MS: (M+H)⁺=482

retention time (HPLC-MS):7.0 min (method B)

7i)(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 70 mg (0.15 mmol)(R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate,

-   51 mg (0.16 mmol) TBTU and 25 μL (0.18 mmol) triethylamine in 1 mL    DMF was stirred for 1 h at RT. Then 25 mg (0.15 mmol)    1-(tetrahydropyran-4-yl)-piperazine was added and the reaction    solution was stirred for 16 h. The reaction solution was purified by    HPLC without any further working up; the fractions containing the    product were combined and lyophilised.

Yield: 36 mg (39% of theory)

ESI-MS: (M+H)⁺=634

retention time (HPLC-MS):5.7 min (method B)

Example 7.1(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-oxy-4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

30 mg (0.17 mmol) MCPBA were added to a solution, cooled to 0° C., of100 mg (0.16 mmol)(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 3 mL chloroform and the reaction solution was stirred for 2 h. Afterthe solvent had been eliminated the residue was taken up in 1 mL DMF andpurified by HPLC; the fractions containing the product were combined andlyophilised.

Yield: 70 mg (69% of theory)

ESI-MS: (M+H)⁺=650

retention time (HPLC-MS):4.1 min (method C)

Example 7.2(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-[4-(4-methyl-tetrahydropyran-4-yl)-piperazin-1-yl]-2-oxo-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

50 mg (0.23 mmol) 1-(4-methyl-tetrahydropyran-4-yl)-piperazine (amineA1, used as the hydrochloride salt) were added to a solution of 90 mg(0.19 mmol) (R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate(Example 7h), 70 mg (0.22 mmol) TBTU and 70 μL (0.50 mmol) triethylaminein 1 mL DMF and the reaction mixture was stirred for 20 h at RT.Semisaturated NaHCO₃ solution was added, the precipitate was filteredoff and dried. The purification was carried out by chromatography (Alox,activity stage II-III, DCM/MeOH 30:1). After the solvent had beeneliminated the residue was triturated with diethyl ether, suctionfiltered and dried.

Yield: 56 mg (44% of theory)

ESI-MS: (M+H)⁺=648

retention time (HPLC-MS):3.1 min (method A)

Example 7.3(R)-1-(4-acetoxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 42 mg (0.07 mmol)(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate(Example 71) in 5 mL acetic anhydride was heated to 50° C. for 2 h. Themixture was evaporated down i.vac. and the residue was purified by HPLC.The fractions containing the product were combined and lyophilised.

Yield: 16 mg (37% of theory)

ESI-MS: (M+H)⁺=676

retention time (HPLC-MS):2.8 min (method A)

Example 7.4(R)-1-(4-formyloxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 25 μL acetic anhydride and 0.5 mL formic acid in 1 mL DCMwas stirred for 2 h at RT (formation of the mixed anhydride). Then 38 mg(0.06 mmol)

(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatewas added (Example 71) and the reaction mixture was stirred overnight atRT. The reaction solution was evaporated down and the residue was againadded to a solution of the mixed anhydride and stirred overnight at RT.The mixture was evaporated down i.vac. and the residue was purified byHPLC. The fractions containing the product were combined andlyophilised.

Yield: 19 mg (45% of theory)

ESI-MS: (M+H)⁺=662

retention time (HPLC-MS):7.2 min (method E)

Example 7.5(R)-1-(4-methoxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

7.5a) (R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A suspension of 1.00 g (1.75 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate(Example 7g) and 150 mg 10% Pd/C in 30 mL MeOH was hydrogenated at RTand 3000 hPa hydrogen pressure until the reaction stopped. The catalystwas suction filtered and the residue was purified by chromatography(silica gel, EtOAc).

Yield: 370 mg (43% of theory)

ESI-MS: (M−H)⁻=494

retention time (HPLC-MS):4.1 min (method A)

7.5b) (R)-2-(4-methoxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

19 μL (0.30 mmol) iodomethane were added to a solution of 100 mg (0.20mmol) (R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 2 mL DMF and the mixture was stirred for 15 min at RT. Then 131 mg(0.40 mmol) Cs₂CO₃ was added and the reaction solution was heated to 50°C. for 3 h. The precipitate was filtered, the filtrate was evaporateddown to 1 mL down and combined with 3 mL water. The precipitate wassuction filtered, dried and further reacted without purification.

Yield: 84 mg (82% of theory)

ESI-MS: (M+H)⁺=510

retention time (HPLC-MS):4.6 min (method A)

7.5c) (R)-2-(4-methoxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 11.3 mg (0.47 mmol) LiOH*H₂O in 8 mL water was added to asolution of 160 mg (0.31 mmol)(R)-2-(4-methoxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 15 mL THF and the reaction mixture was stirred for 1 h at RT. Themixture was evaporated down i.vac., the residue was taken up in 150 mLwater and acidified with 2 M HCl. The precipitate formed was separatedoff and dried.

Yield: 138 mg (89% of theory)

ESI-MS: (M−H)⁻=494

retention time (HPLC-MS):4.0 min (method A)

7.5d)(R)-1-(4-methoxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 40 mg (0.08 mmol)(R)-2-(4-methoxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate,

-   29 mg (0.09 mmol) TBTU and 14 μL (0.10 mmol) triethylamine in 1 mL    DMF was stirred for 1 h at RT. Then 15 mg (0.09 mmol)    1-(tetrahydropyran-4-yl)-piperazine was added and the mixture was    stirred for a further 16 h at RT. The reaction solution was purified    by HPLC without any further working up; the fractions containing the    product were combined and lyophilised.

Yield: 19 mg (37% of theory)

ESI-MS: (M+H)⁺=648

retention time (HPLC-MS):3.3 min (method A)

Example 8(R)-1-(3-ethyl-4-hydroxy-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

8a) 2-ethyl-6-methyl-phenol

19.4 mL (0.23 mmol) concentrated HCl and a solution of 16.1 g (0.23mmol) sodium nitrite in water (approx. 70 mL) were added to a solutionof 30 g (222 mmol) 2-ethyl-6-methyl-aniline in 135 mL EtOH at 0° C. andthe mixture was stirred for 15 min. This mixture was added at 45° C. toa solution of 10.5 mL concentrated H₂SO₄ in 300 mL water and at the endof the addition heated to 70° C. The aqueous phase was cooled to RT andexhaustively extracted with EtOAc. The combined organic phases wereextracted with 1 M NaOH solution. The aqueous phase was washed with DCM,acidified to pH 1 with 4 N HCl solution and extracted with DCM. Theorganic phase was washed with saturated NaCl solution, dried over Na₂SO₄and evaporated down i.vac. The crude product was used in the nextreaction step without any further purification.

Yield: 12.0 g (40% of theoretical)

8b) 4-bromo-2-ethyl-6-methyl-phenol

At RT a solution of 12.7 mL (247 mmol) bromine in 10 mL chloroform wasadded dropwise to a solution of 33.6 g (247 mmol)2-ethyl-6-methyl-phenol in 350 mL chloroform and the mixture was stirredfor 2 h. The reaction mixture was combined with an aqueous NaHSO₃solution and stirred for 20 min. The phases were separated and theorganic phase was washed with saturated NaCl solution, dried over Na₂SO₄and evaporated down i.vac. Column chromatography (silica gel, Cyc/EtOAc9:1) yielded the product.

Yield: 39.8 g (75% of theoretical)

ESI-MS: (M+H)⁺=214/216 (Br)

retention time (HPLC-MS):6.3 min (method D)

8c) 2-benzyloxy-5-bromo-1-ethyl-3-methyl-benzene

A suspension of 39.8 g (185 mmol) 4-bromo-2-ethyl-6-methyl-phenol, 63.9g (0.46 mmol) K₂CO₃ and 22.0 mL (185 mmol) benzylbromide in 450 mLacetonitrile was refluxed for 3 h, cooled to RT and evaporated downi.vac. The residue was combined with EtOAc, the organic phase was washedwith water and saturated NaCl solution, dried over Na₂SO₄ and evaporateddown i.vac.

Yield: 54.5 g (96% of theoretical)

ESI-MS: (M+H)⁺=304/306 (Br)

retention time (HPLC-MS):9.4 min (method D)

8d) methyl(Z,E)-2-acetylamino-3-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-acrylate

Prepared analogously to Example 7b from 50.4 g (165.1 mmol)2-benzyloxy-5-bromo-1-ethyl-3-methyl-benzene and 28.9 g (198.2 mmol)methyl 2-acetylamino-acrylate.

Yield: 41.0 g (68% of theoretical)

ESI-MS: (M+H)⁺=368

retention time (HPLC-MS):4.5 min (method A)

8e) 3-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-2-oxo-propionic acid

200 mL 4 M HCl were added to a solution of 41.0 g (111.6 mmol) methyl(Z,E)-2-acetylamino-3-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-acrylate in300 mL 1,4-dioxane and the reaction solution was heated to 130° C. (bathtemperature) for 7 h. The organic phase was separated off while hot,evaporated down i.vac. and the residue obtained was recrystallised fromtoluene.

Yield: 9.6 g (28% of theoretical)

ESI-MS: (M+H)⁺=312

retention time (HPLC-MS):4.1 min (method A)

8f) (R)-3-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-2-hydroxy-propionic acid

Under an argon atmosphere a solution of 9.59 g (30.7 mmol)3-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-2-oxo-propionic acid in 25 mLTHF was combined with 4.26 mL (31.0 mmol) triethylamine, stirred for 5min and cooled to −30° C. (internal temperature). A solution of 19.7 g(61.0 mmol) (1R)-B-chlorodiisopinocampheylborane in 35 mL THF was addeddropwise and after the end of the addition the reaction solution wasstirred for 30 min without cooling. 15 mL 4 N NaOH was added(temperature rose to 20° C.), the mixture was stirred for 5 min, cooledto 0° C., combined with 50 mL MTBE and stirred for 20 min. The organicphase was separated off and dried over Na₂SO₄. After the desiccant andsolvent had been eliminated the residue was further reacted withoutpurification.

Yield: 10.3 g (100% of theoretical)

ESI-MS: (M−H)⁻=313

retention time (HPLC-MS):4.2 min (method A)

8g) methyl(R)-3-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-2-hydroxy-propionate

Prepared analogously to Example 7e from 10.3 g (30.7 mmol)(R)-3-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-2-hydroxy-propionic acid and4.71 mL (64.5 mmol) thionyl chloride. The crude product obtained wasfurther reacted without purification.

8h) (R)-2-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 7.12 g (34.3 mmol) 4-nitrophenyl-chloroformate in 30 mLTHF was added within 10 min to a solution of 75 mL pyridine heated to60° C. (bath temperature), the mixture was stirred for 10 min and then asolution of 10.0 g of the crude product from Example 8g in 50 mLpyridine was added dropwise. The mixture was stirred for 1 h, combinedwith 6.72 g (27.4 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one and the bathtemperature was increased to 100° C. (2 h). The precipitate formed wasfiltered, the filtrate was evaporated down i.vac., the residue wascombined with 150 mL EtOAc, the organic phase was washed twice with 50mL 1 M KHSO₄ solution and ten times with 50 mL 15% K₂CO₃ solution anddried over Na₂SO₄. After the desiccant and solvent had been eliminatedthe residue was purified by chromatography (silica gel, EtOAc/Cyc 2:1).

Yield: 2.28 g (14% of theoretical)

ESI-MS: (M+H)⁺=600

retention time (HPLC-MS):5.4 min (method A)

8i) (R)-2-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 50 mg (2.09 mmol) LiOH in 5 mL water was added to asolution of 800 mg (1.33 mmol)(R)-2-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-1-methoxy-carbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 15 mL THF and the reaction mixture was stirred for 1 h at RT. Themixture was evaporated down i.vac., the residue was taken up in 50 mLwater and 2 M HCl was added until an acidic reaction was obtained. Theprecipitate formed was filtered off, washed with water and dried.Further purification was carried out by decocting with 150 mL water,filtration and drying again.

Yield: quantitative

ESI-MS: (M+H)⁺=586

retention time (HPLC-MS):4.8 min (method A)

8k (R)-1-carboxy-2-(3-ethyl-4-hydroxy-5-methyl-phenyl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

810 mg (1.38 mmol)(R)-2-(4-benzyloxy-3-ethyl-5-methyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 25 mL MeOH were combined with 80 mg 10% Pd/C and hydrogenated at RTand 3000 hPa hydrogen until the reaction stopped. The catalyst wassuction filtered and the solvent was evaporated down i.vac. The residuewas triturated with DIPE, suction filtered and dried.

Yield: 639 mg (93% of theory)

ESI-MS: (M+H)⁺=496

retention time (HPLC-MS):3.7 min (method A)

8i)(R)-1-(3-ethyl-4-hydroxy-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

30 mg (0.18 mmol) 1-(tetrahydropyran-4-yl)-piperazine were added at RTto a solution of 80 mg (0.16 mmol)(R)-1-carboxy-2-(3-ethyl-4-hydroxy-5-methyl-phenyl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate,57 mg (0.18 mmol) TBTU and 28 μL (0.20 mmol) triethylamine in 1 mL DMFand the reaction mixture was stirred for 2 h. The reaction solution waspurified by HPLC without any further working up, the fractionscontaining the product were combined and lyophilised.

Yield: 75 mg (72% of theory)

ESI-MS: (M+H)⁺=648

retention time (HPLC-MS):3.2 min (method A)

Example 9(R)-1-(4-hydroxy-3-methoxy-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

9a) 4-bromo-2-methoxy-6-methyl-phenol

A solution of 56.2 g (0.32 mol) N-bromosuccinimide in 1700 mL AcOH wasadded dropwise within 5.5 h to a solution of 42.3 g (0.31 mol)2-methoxy-6-methyl-phenol in 450 mL AcOH and the mixture was stirred for16 h at RT. The reaction mixture was evaporated down i.vac. and theresidue was taken up in DCM. The organic phase was washed with 5% NaHCO₃and saturated NaCl solution, dried over Na₂SO₄ and evaporated downi.vac. The red oil was used in the next reaction step without anyfurther purification.

Yield: 65.9 g (66% of theoretical)

R_(f)=0.32 (silica gel, hexane/EtOAc 4:1)

retention time (HPLC-MS):11.1 min (method D)

9b) 2-benzyloxy-5-bromo-1-methoxy-3-methyl-benzene

45.7 g (0.33 mol) K₂CO₃ and a solution of 40.3 mL (0.33 mol)benzylbromide were added at RT to a solution of 65.9 g (0.26 mol)4-bromo-2-methoxy-6-methyl-phenol in 330 mL DMF and the mixture wasstirred for 18 h at RT. The mixture was filtered, evaporated down i.vac.and the residue was taken up in diethyl ether. The organic phase waswashed with water, 5% Na₂CO₃ and NaCl solution, dried over Na₂SO₄ andevaporated down i.vac. The crude product was used in the next reactionstep without any further purification.

Yield: 92.2 g (81% of theoretical)

R_(f)=0.56 (silica gel, hexane/EtOAc 4:1)

retention time (HPLC-MS):16.3 min (method D)

9c) 4-benzyloxy-3-methoxy-5-methyl-benzaldehyde

96 mL (240 mmol) n-butyllithium (2.5 M in hexane) were added dropwise at−75° C. to a solution of 61.2 g (119.5 mmol)2-benzyloxy-5-bromo-1-methoxy-3-methyl-benzene in 240 mL THF and themixture was stirred for 15 min at −75° C. A solution of 31 mL (402 mmol)DMF in 30 mL THF was added dropwise, the mixture was heated to 0° C. andstirred for a further 2 h. The reaction was combined with saturatedNH₄Cl solution, diluted with 150 mL water and the phases were separated.The aqueous phase was exhaustively extracted with diethyl ether. Thecombined organic phases were washed with saturated NaCl solution, driedover Na₂SO₄ and evaporated down i.vac. Column chromatography (silicagel, hexane/EtOAc 85:15) yielded the product as a yellow oil.

Yield: 27.1 g (88% of theoretical)

R_(f)=0.32 (silica gel, hexane/EtOAc 4:1)

retention time (HPLC-MS):13.3 min (method D)

9d) 2-acetylamino-3-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-acrylic acid

A suspension of 27.0 g (105.4 mmol)4-benzyloxy-3-methoxy-5-methyl-benzaldehyde, 18.5 g (158.0 mmol)N-acetylglycine and 12.96 g (158.0 mmol) NaOAc in 120 mL aceticanhydride was heated to 115° C. under nitrogen for 3.5 h. At 100° C. 60mL water were slowly added dropwise and the mixture was stirred for 1 h.The reaction mixture was cooled to RT, poured into water and the aqueousphase was exhaustively extracted with EtOAc. The combined organic phaseswere washed with saturated NaCl solution, dried over Na₂SO₄ andevaporated down i.vac. The residue was triturated with isopropanol, thesolid obtained was washed with isopropanol, diethyl ether and a littleacetone and dried i.vac. at 45° C.

Yield: 21.2 g (57% of theoretical)

R_(f)=0.24 (silica gel, hexane/EtOAc 4:1)

retention time (HPLC-MS):9.4 min (method D)

9e) 3-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-2-oxo-propionic acid

The product was obtained analogously to Example 7c starting from 20.0 g(56.3 mmol)2-acetylamino-3-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-acrylic acid.The crude product was used in the next reaction step without any furtherpurification.

Yield: 15.6 g (53% of theoretical) retention time (HPLC-MS):11.9 min(method D) 9f)(R)-3-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-2-hydroxy-propionic acid

The product was prepared analogously to Example 7d starting from 16.0 g(50.90 mmol) 3-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-2-oxo-propionicacid.

Yield: 7.63 g (47% of theoretical) retention time (HPLC-MS):9.8 min(method D)

9g) methyl(R)-3-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-2-hydroxy-propionate

The product was prepared analogously to Example 7e starting from 7.6 g(24.02 mmol)(R)-3-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-2-hydroxy-propionic acid.

Yield: 6.84 g (86% of theoretical)

retention time (HPLC-MS):11.7 min (method D)

9h)(R)-2-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

The product was prepared analogously to Example 7f starting from 6.8 g(20.6 mmol) methyl(R)-3-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-2-hydroxy-propionate inacetonitrile.

Yield: 8.16 g (66% of theoretical)

ESI-MS: (M+H)⁺=602

retention time (HPLC-MS):14.1 min (method D)

9i) (R)-2-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

The product was prepared analogously to Example 7g starting from 8.16 g(13.65 mmol)(R)-2-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate.

Yield: 7.83 g (98% of theoretical)

ESI-MS: (M+H)⁺=588

retention time (HPLC-MS):12.2 min (method D)

9k) (R)-1-carboxy-2-(4-hydroxy-3-methoxy-5-methyl-phenyl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

The product was prepared analogously to Example 7h starting from 7.80 g(13.27 mmol)(R)-2-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate.

Yield: 5.33 g (80% of theoretical)

ESI-MS: (M+H)⁺=498

retention time (HPLC-MS):8.4 min (method D)

9i)(R)-1-(4-hydroxy-3-methoxy-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-Piperidine-1-carboxylate

84 mg (0.22 mmol) HATU was added at RT under nitrogen to a solution of100 mg (0.20 mmol)(R)-1-carboxy-2-(4-hydroxy-3-methoxy-5-methyl-phenyl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate,110 μL (0.64 mmol) ethyldiisopropylamine and 59 mg (0.24 mmol)1-(tetrahydropyran-4-yl)-piperazine (used as the bis-hydrochloride salt)in 5 mL DMF and the mixture was stirred for 3 h. The reaction mixturewas evaporated down i.vac. at 50° C. and the crude product was purifiedby HPLC-MS; the fractions containing the product were combined andlyophilised.

Yield: 112 mg (73% of theoretical)

ESI-MS: (M+H)⁺=650

retention time (HPLC-MS):3.6 min (method D)

Example 10{(R)-1-(3-chloro-4-hydroxy-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl}-amide4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

10a) 2-benzyloxy-5-bromo-1-chloro-3-methyl-benzene

A mixture of 10.2 g (46.0 mmol) 4-bromo-2-chloro-6-methyl-phenol, 7.0 mL(57.6 mmol) benzylbromide, 30.0 g (217.1 mmol) K₂CO₃ and 130 mL DMF wasstirred overnight at RT. After the reaction had ended the insolublematter was filtered off, the filtrate was evaporated down i.vac.,combined with water and exhaustively extracted with EtOAc. The organicphases were combined, dried over Na₂SO₄ and evaporated down i.vac.

Yield: 14.0 g (98% of theory)

R_(f)=0.91 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

10b) methyl2-acetylamino-3-(4-benzyloxy-3-chloro-5-methyl-phenyl)-acrylate

Prepared analogously to Example 7b from 28.0 g (89.8 mmol)2-benzyloxy-5-bromo-1-chloro-3-methyl-benzene, 15.0 g (102.7 mmol)methyl 2-acetylamino-acrylate, 260 mL triethylamine, 400 mLacetonitrile, 4.4 g (14.0 mmol) tri-o-tolyl-phosphane and 3.2 g (14.2mmol) Pd(OAc)₂.

Yield: 12.5 g (37% of theory)

ESI-MS: (M+H)⁺=374/376 (Cl)

R_(f)=0.67 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

10c) methyl2-acetylamino-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-propionate

A mixture of 7.40 g (19.8 mmol) methyl2-acetylamino-3-(4-benzyloxy-3-chloro-5-methyl-phenyl)-acrylate, 300 mLMeOH and 800 mg Raney nickel was shaken at RT and 3000 hPa hydrogenpressure for 7 h. After the reaction had ended the catalyst was filteredoff and the remainder was evaporated down i.vac.

Yield: 5.6 g (99% of theory)

ESI-MS: (M+H)⁺=286/288 (Cl)

retention time (HPLC-MS):3.0 min (method A)

10d) methyl(R)-2-acetylamino-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-propionate

6.0 mL Alcalase 2.4 L FG (Novozymes NS; DK 2880 Bagsvaerd) was added toa warm solution (37° C.) of 7.2 g (40.4 mmol) Na₂HPO₄ dihydrate in 100mL water and by the addition of NaH₂PO₄ dihydrate the pH was adjusted to7.5. Then a solution of 5.5 g (19.2 mmol) methyl2-acetylamino-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-propionate in 50 mLacetone was added dropwise with stirring at 37° C. The pH value of thereaction mixture was kept constantly in the range from pH 7.4-7.6 by theaddition of 1 M NaOH. After the addition had ended the mixture wasstirred for 4 h at 37° C. After cooling to RT the reaction mixture wasexhaustively extracted with MTBE, the combined organic extracts werewashed with 15% K₂CO₃ solution and dried over Na₂SO₄. After thedesiccant and solvent had been eliminated the crude product (1.6 g) wasfurther reacted without purification.

ESI-MS: (M+H)⁺=286/288 (Cl)

10e) methyl(R)-2-amino-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-propionate

A mixture of 1.5 g of the above crude product and 8.75 mL of 4 M HCl wasrefluxed for 5 h. The mixture was evaporated down i.vac., the residuewas taken up in water and made alkaline by the addition of K₂CO₃solution. The aqueous phase was acidified by the addition of 4 M HCl,exhaustively extracted with EtOAc, the combined organic extracts weredried and evaporated down i.vac. The residue was combined withmethanolic HCl and stirred overnight at RT. The reaction mixture wasevaporated down i.vac., the residue was taken up in 15% K₂CO₃ solutionand exhaustively extracted with EtOAc. The combined organic phases weredried and evaporated down i.vac.

Yield: 0.50 g (39% of theory)

ESI-MS: (M+H)⁺=244/246 (Cl)

R_(f)=0.59 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

10f) methyl(R)-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-2-{[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carbonyl]-amino}-propionate

0.39 g (2.4 mmol) CDT were added to a solution of 0.5 g (2.1 mmol)methyl (R)-2-amino-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-propionate in20 mL THF cooled in the ice bath and the reaction mixture was stirredfor 1 h while cooling with ice and for 1 h at RT. Then 0.54 g (2.2 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one were addedand the reaction mixture was refluxed for 3 h. The mixture wasevaporated down i.vac., the residue was combined with 1 M KHSO₄solution, the precipitate formed was suction filtered and dried.

Yield: 1.0 g (95% of theory)

ESI-MS: (M+H)⁺=513/515 (Cl)

R_(f)=0.55 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

10g)(R)-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-2-{[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carbonyl]-amino}-propionicacid

A solution of 0.07 g (3.0 mmol) LiOH in 3 mL water was added to asolution of 1.0 g (1.94 mmol) methyl(R)-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-2-{[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carbonyl]-amino}-propionatein 15 mL THF and the reaction mixture was stirred overnight at RT. TheTHF was eliminated i.vac., 100 mL of water were added and the mixturewas acidified with 2 M HCl. The precipitated product was suctionfiltered, washed with 50 mL water and dried at 60° C. in the dryingcupboard.

Yield: 0.9 g (93% of theory)

ESI-MS: (M+H)⁺=501/503 (Cl)

R_(f)=0.08 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

10h){(R)-1-(3-chloro-4-hydroxy-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl}-amide4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 71 from 70 mg (0.14 mmol)(R)-3-(3-chloro-4-hydroxy-5-methyl-phenyl)-2-{[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carbonyl]-amino}-propionicacid and 27.4 mg (0.16 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 52 mg (57% of theory)

ESI-MS: (M+H)⁺=653/655 (Cl)

retention time (HPLC-MS):2.5 min (method A)

Example 11(R)-1-(3-chloro-4-hydroxy-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

11a) 3-(4-benzyloxy-3-chloro-5-methyl-phenyl)-2-oxo-propionic acid

Prepared analogously to Example 7c from 18.4 g (49.2 mmol) methyl2-acetylamino-3-(4-benzyloxy-3-chloro-5-methyl-phenyl)-acrylate (Example10b) and 75 mL 4 M HCl.

Yield: 15.5 g (99% of theory)

ESI-MS: (M+H)⁺=317/319 (Cl)

R_(f)=0.20 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

11b) methyl(R)-3-(4-benzyloxy-3-chloro-5-methyl-phenyl)-2-hydroxy-propionate

(R)-3-(4-benzyloxy-3-chloro-5-methyl-phenyl)-2-hydroxy-propionic acidwas prepared analogously to Example 7d from 15.5 g (48.6 mmol)3-(4-benzyloxy-3-chloro-5-methyl-phenyl)-2-oxo-propionic acid and 27.6 g(86.1 mmol) (1R)-B-chlorodiisopinocampheylborane. The crude product wascombined with 150 mL methanolic HCl (1.25 M) and stirred overnight atRT. The reaction solution was evaporated down i.vac., the residue wascombined with 70 mL 15% K₂CO₃ solution and extracted twice with 50 mLEtOAc. The combined organic extracts were dried, filtered and evaporateddown i.vac.

Yield: 7.0 g (43% of theory)

ESI-MS: (M+NH₄)⁺=352/354 (Cl)

R_(f)=0.87 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

11c) (R)-2-(4-benzyloxy-3-chloro-5-methyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

(R)-2-(4-benzyloxy-3-chloro-5-methyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatewas prepared analogously to Example 7f from 7.0 g (20.9 mmol) methyl(R)-3-(4-benzyloxy-3-chloro-5-methyl-phenyl)-2-hydroxy-propionate and5.2 g (21.2 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one. The crudeproduct was dissolved in 150 mL THF and combined with a solution of 0.50g (20.6 mmol) LiOH in 50 mL water. The reaction mixture was stirredovernight at RT, combined with water and the organic solvent waseliminated i.vac. The aqueous phase was washed twice with EtOAc andacidified with 21 mL 1 M HCl. The resulting oil was exhaustivelyextracted with EtOAc. The combined organic extracts were dried, filteredand evaporated down i.vac.

Yield: 3.3 g (26% of theory)

ESI-MS: (M+H)⁺=592/594 (Cl)

R_(f)=0.35 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

11d) (R)-1-carboxy-2-(3-chloro-4-hydroxy-5-methyl-phenyl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

200 mg 5% Rh on aluminium oxide were added to a solution of 800 mg (1.35mmol) (R)-2-(4-benzyloxy-3-chloro-5-methyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 20 mL THF and the suspension was hydrogenated for 12 h at 40° C. and3000 hPa hydrogen pressure. To complete the reaction the mixture wascombined with 5 mL DCM/MeOH (1:1) and hydrogenated for a further 20 h at40° C. and 3000 hPa. The catalyst was filtered off, the filtrate wasevaporated down, the residue was triturated with DIPE, suction filteredand dried.

Yield: 639 mg (94% of theory)

ESI-MS: (M+H)⁺=502/504 (Cl)

retention time (HPLC-MS):3.6 min (method A)

11e)(R)-1-(3-chloro-4-hydroxy-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 71 from 80 mg (0.16 mmol)(R)-1-carboxy-2-(3-chloro-4-hydroxy-5-methyl-phenyl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)piperidine-1-carboxylateand 29.8 mg (0.18 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 13 mg (12% of theory)

ESI-MS: (M+H)⁺=654/656 (Cl)

retention time (HPLC-MS):3.2 min (method A)

Example 11.1(R)-1-(4-hydroxy-3-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

11.1a)(R)-1-(4-benzyloxy-3-chloro-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 150 mg (0.25 mmol)(R)-2-(4-benzyloxy-3-chloro-5-methyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate,89 mg (0.28 mmol) TBTU and 44 μL (0.32 mmol) triethylamine in 1.5 mL DMFwas stirred for 1 h at RT. Then 47 mg (0.28 mmol)1-(tetrahydropyran-4-yl)-piperazine were added and the reaction solutionwas stirred for 2 h. The reaction solution was purified by HPLC withoutany further working up; the fractions containing the product werecombined and lyophilised.

Yield: 108 mg (57% of theory)

ESI-MS: (M+H)⁺=744/746 (Cl)

retention time (HPLC-MS):4.1 min (method A)

11.1b)(R)-1-(4-hydroxy-3-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A suspension of 108 mg (0.15 mmol)(R)-1-(4-benzyloxy-3-chloro-5-methyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 30 mg 10% Pd/C in 10 mL MeOH and 0.5 mL triethylamine washydrogenated at RT and 3000 hPa hydrogen pressure until the theoreticalamount of hydrogen had been taken up. The catalyst was filtered off, theresidue was dissolved in 1 mL DMF and purified by HPLC. The fractionscontaining the product were combined and lyophilised.

Yield: 39 mg (43% of theory)

ESI-MS: (M+H)⁺=620

retention time (HPLC-MS):3.0 min (method A)

Example 12(S)-2-(3-chloro-4-hydroxy-5-methyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

12a) 3-chloro-4-hydroxy-5-methyl-benzaldehyde

79.2 mL of a 2.5 M solution of n-butyllithium in n-hexane was addeddropwise within 30 min to a mixture, cooled to −70° C., of 20.0 g (90.3mmol) 4-bromo-2-chloro-6-methyl-phenol and 250 mL of THF. The reactionmixture was stirred for 2 h and then combined dropwise with 28.47 mL(370 mmol) DMF and heated to RT overnight. Then 150 mL 2 M HCl was addeddropwise to the reaction solution while cooling with an ice bath, themixture was stirred for 15 min and the pH was adjusted to 9-10 by theaddition of saturated NaHCO₃ solution. The organic phase was separatedoff and discarded and the aqueous phase was extracted twice with EtOAc.The combined organic phases were dried, filtered and evaporated downi.vac.

Yield: 15.1 g (98% of theory)

ESI-MS: (M−H)⁻=169/171 (Cl)

R_(f)=0.93 (silica gel, EtOAc)

12b) 1-methyl2-[1-(3-chloro-4-hydroxy-5-methyl-phenyl)-meth-(Z)-ylidene]-succinate

69.5 g (177.0 mmol) 1-methyl 2-(triphenyl-λ⁵-phosphanylidene)-succinatewere added to a solution of 15.0 g (87.9 mmol)3-chloro-4-hydroxy-5-methyl-benzaldehyde in 250 mL THF and the reactionmixture was heated to 40° C. for 120 h. The mixture was evaporated downi.vac., water and EtOAc were combined with the residue, the organicphase was separated off, washed with water and extracted three timeswith 200 mL 5% K₂CO₃ solution each time. The combined aqueous phaseswere acidified with semiconcentrated HCl and the oily precipitate wasextracted twice with 250 mL EtOAc each time. The combined organic phaseswere dried, filtered and evaporated down i.vac.

Yield: 11.1 g (44% of theory)

EI: (M−H)⁻=283/285 (Cl)

R_(f)=0.70 (silica gel, EtOAc)

12c) 1-methyl (S)-2-(3-chloro-4-hydroxy-5-methyl-benzyl)-succinate

Under an argon atmosphere 450 mg(−)-1,2-bis((2R,5R)-2,5-diethyl-phospholano)benzene(cyclooctadiene)rhodium(I)tetrafluoroboratewere added to a solution of 11.0 g (38.6 mmol) 1-methyl2-[1-(3-chloro-4-hydroxy-5-methyl-phenyl)-meth-(Z)-ylidene]-succinate in150 mL degassed MeOH and 11.0 mL triethylamine and the reaction mixturewas hydrogenated at 3447 hPa hydrogen pressure for 8 h. Then thereaction solution was evaporated down i.vac., the residue was dissolvedin 100 mL EtOAc, washed twice with 2 M HCl and exhaustively extractedwith 15% K₂CO₃ solution. The aqueous phase was acidified withconcentrated HCl, exhaustively extracted with EtOAc and the organicphase was dried over Na₂SO₄. After the desiccant and solvent had beeneliminated the desired product was obtained.

Yield: 11.0 g (99% of theory)

ESI-MS: (M−H)⁻=285/287 (Cl)

retention time (HPLC-MS):3.3 min (method A)

12d) methyl(S)-2-(3-chloro-4-hydroxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoate

6.6 g (27.0 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one was added toa mixture of 7.0 g (24.4 mmol) 1-methyl(S)-2-(3-chloro-4-hydroxy-5-methyl-benzyl)-succinate, 8.7 g (27.0 mmol)TBTU, 4.65 mL (27 mmol) ethyldiisopropylamine, 100 mL THF and 10 mL DMFand the reaction mixture was stirred overnight at RT. The reactionsolution was evaporated down i.vac., the residue was taken up in DCM,the organic phase was washed with 15% Na₂CO₃ solution and dried overNa₂SO₄. After the desiccant and solvent had been eliminated the desiredproduct was obtained.

Yield: 12.1 g (96% of theory)

ESI-MS: (M+H)⁺=514/516 (Cl)

R_(f)=0.49 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

12e)(S)-2-(3-chloro-4-hydroxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid

Prepared analogously to Example 7g from 12.1 g (23.5 mmol) methyl(S)-2-(3-chloro-4-hydroxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoateand 848 mg (34.4 mmol) LiOH.

Yield: 9.7 g (82% of theory)

ESI-MS: (M+H)⁺=500/502 (Cl)

R_(f)=0.31 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

12f)(S)-2-(3-chloro-4-hydroxy-5-methyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

Prepared analogously to Example 71 from 100 mg (0.20 mmol)(S)-2-(3-chloro-4-hydroxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid and 37.5 mg (0.22 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 33 mg (25% of theory)

ESI-MS: (M+H)⁺=652/654 (Cl)

retention time (HPLC-MS):3.3 min (method B)

Example 13(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(5-oxo-3-phenyl-4,5-dihydro-1,2,4-triazol-1-yl)-piperidine-1-carboxylate

13a) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(5-oxo-3-phenyl-4,5-dihydro-1,2,4-triazol-1-yl)-piperidine-1-carboxylate

Prepared analogously to Example 7f from 5.0 g (15.9 mmol) methyl(R)-3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-hydroxy-propionate (Example7e) and 5.98 g (15.9 mmol)5-phenyl-2-piperidin-4-yl-2,4-dihydro-1,2,4-triazol-3-one (purity 65%).

Yield: 4.96 g (53% of theory)

ESI-MS: (M+H)⁺=585

retention time (HPLC-MS):5.0 min (method A)

13b) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(5-oxo-3-phenyl-4,5-dihydro-1,2,4-triazol-1-yl)-piperidine-1-carboxylate

A solution of 310 mg (12.93 mmol) LiOH in 30 mL water was added to asolution of 4.96 g (8.48 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxy-carbonyl-ethyl4-(5-oxo-3-phenyl-4,5-dihydro-1,2,4-triazol-1-yl)-piperidine-1-carboxylatein 50 mL THF and the reaction mixture was stirred for 7 h at RT. Thesolution was stored overnight at −18° C. and after heating to RT afurther 310 mg LiOH were added thereto to complete the reaction. After 1h the reaction solution was evaporated down i.vac., the residue wastaken up in 150 mL water and acidified with 1 M HCl. The precipitate wasfiltered off and dried at 40° C.

Yield: 4.75 g (98% of theory)

ESI-MS: (M+H)⁺=571

retention time (HPLC-MS):4.3 min (method A)

13c) (R)-1-carboxy-2-(4-hydroxy-3,5-dimethyl-phenyl)-ethyl4-(5-oxo-3-phenyl-4,5-dihydro-1,2,4-triazol-1-yl)-piperidine-1-carboxylate

A solution of 2.50 g (4.38 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(5-oxo-3-phenyl-4,5-dihydro-1,2,4-triazol-1-yl)-piperidine-1-carboxylatein 50 mL DCM was combined with 250 mg 10% Pd/C and hydrogenated at RTunder a hydrogen pressure of 3000 hP for 4.5 h. To complete the reactiona further 250 mg catalyst were added, the mixture was hydrogenated for12 h at 40° C., combined with 25 mL THF and 250 mg catalyst andhydrogenated for a further 12 h at 40° C. The catalyst was suctionfiltered and the filtrate was evaporated down i.vac. The residue wasstirred with DIPE, suction filtered and dried.

Yield: 1.87 g (89% of theory)

ESI-MS: (M−H)⁻=479

retention time (HPLC-MS):3.5 min (method A)

13d)(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(5-oxo-3-phenyl-4,5-dihydro-1,2,4-triazol-1-yl)-piperidine-1-carboxylate

A solution of 100 mg (0.21 mmol)(R)-1-carboxy-2-(4-hydroxy-3,5-dimethyl-phenyl)-ethyl4-(5-oxo-3-phenyl-4,5-dihydro-1,2,4-triazol-1-yl)-piperidine-1-carboxylate,74 mg (0.23 mmol) TBTU and 35 μL (0.26 mmol) triethylamine in 1 mL DMFwas stirred for 1 h at RT. Then 35 mg (0.21 mmol)1-(tetrahydropyran-4-yl)-piperazine was added and the reaction mixturewas stirred for a further 5 h at RT. The reaction solution was purifiedby HPLC without any further working up.

The fractions containing the product were combined and lyophilised.

Yield: 39 mg (30% of theory)

ESI-MS: (M+H)⁺=633

retention time (HPLC-MS):2.9 min (method A)

Example 14(R)-1-(4-amino-3-methyl-5-trifluoromethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

14a) 2-methyl-6-trifluoromethyl-phenylamine

A suspension of 50g (0.24 mol)1-methyl-2-nitro-3-trifluoromethyl-benzene and 4.4 g 10% Pd/C in 300 mLMeOH was hydrogenated at RT and 3000 hPa hydrogen pressure until thetheoretical amount of hydrogen had been taken up. The catalyst wasfiltered off, washed with MeOH and the filtrate was evaporated down. Thecrude product was further reacted without purification.

Yield: quantitative

ESI-MS: (M+H)⁺=176

14b) 4-bromo-2-methyl-6-trifluoromethyl-phenylamine

Under a nitrogen atmosphere a solution of 11.0 mL (214 mmol) bromine in100 mL chloroform was added dropwise to a solution of 35.8 g (204 mmol)2-methyl-6-trifluoromethyl-phenylamine in 350 mL chloroform and afterthe end of the addition the reaction mixture was stirred for 3 h at RT.Saturated NaHCO₃ solution was added with stirring, the mixture wasstirred for a further 20 min at RT, the organic phase was separated offand dried over Na₂SO₄. After the desiccant and solvent had beeneliminated the product was obtained as an oil, which was further reactedwithout purification.

Yield: 47.0 g (52% of theory)

EI-MS: (M)⁺=253/255 (Br)

14c) methyl(Z,E)-2-acetylamino-3-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-acrylate

Under a nitrogen atmosphere 5.2 g (23.2 mmol) Pd(OAc)₂ and 7.2 g (22.9mmol) tri-o-tolyl-phosphane were added to a solution of 37.2 g (146mmol) 4-bromo-2-methyl-6-trifluoromethyl-phenylamine and 24.5 g (168mmol) methyl 2-acetylamino-acrylate in 700 mL acetonitrile and 440 mLtriethylamine and the reaction mixture was stirred for 18 h at a bathtemperature of 80° C. After cooling the precipitate formed was suctionfiltered, the filtrate was evaporated to dryness and the residue wascombined with 100 mL water and 50 mL EtOAc. The precipitate was suctionfiltered and dried at 50° C.

Yield: 21.6 g (47% of theory)

ESI-MS: (M+H)⁺=317

R_(f)=0.41 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

14d) 3-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-2-oxo-propionic acid

A solution of 21.6 g (68.3 mmol) methyl(Z,E)-2-acetylamino-3-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-acrylatein 200 mL 1,4-dioxane and 100 mL 4 M HCl was refluxed for 5 h. The1,4-dioxane was eliminated i.vac., the precipitate was filtered off,washed with water and dried at 50° C.

Yield: 11.6 g (65% of theory)

ESI-MS: (M−H)⁻=260

R_(f)=0.11 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

14e)(R)-3-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-2-hydroxy-propionicacid

Under a nitrogen atmosphere a solution of 24.5 g (76.3 mmol)(1R)-B-chlorodiisopinocampheylborane in 100 mL THF was added dropwisewithin 15 min to a solution, cooled to −35° C., of 11.6 g (44.4 mmol)3-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-2-oxo-propionic acid and8.1 mL (58.3 mmol) triethylamine in 200 mL THF and the reaction solutionwas stirred overnight at RT. Then at RT the reaction solution wascarefully made alkaline with 23 mL 4 M NaOH, combined with 200 mL MTBEand 150 mL water and stirred for 1 h. The aqueous phase was separatedoff, the organic phase was extracted twice with 50 mL water and thecombined aqueous extracts were acidified with 4 M HCl. The mixture wasextracted three times with 100 mL EtOAc and the combined organic phaseswere dried over MgSO₄. After the desiccant and solvent had beeneliminated the residue was further reacted without purification.

Yield: 8.4 g (72% of theory)

ESI-MS: (M+H)⁺=264

R_(f)=0.11 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

14f) methyl(R)-3-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-2-hydroxy-propionate

A solution of 8.4 g (31.9 mmol)(R)-3-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-2-hydroxy-propionicacid in 100 mL methanolic HCl (1.3 M) was stirred for 3 h at RT. Themixture was evaporated down i.vac., the residue was combined with 150 mLof 15% K₂CO₃ solution, extracted three times with 100 mL EtOAc and thecombined organic phases were dried over MgSO₄. After the desiccant andsolvent had been eliminated the residue was further reacted withoutpurification.

Yield: 6.1 g (69% of theory)

ESI-MS: (M+H)⁺=278

R_(f)=0.77 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

14g)(R)-2-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere 4.7 g (23.3 mmol) 4-nitrophenylchloroformate in 30 mL THF were metered into 60 mL pyridine at a bathtemperature of 60° C. within 10 min, the mixture was stirred for 5 min,then 6.1 g (22.0 mmol) methyl(R)-3-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-2-hydroxy-propionatein 40 mL pyridine were added and the reaction mixture was stirred for2.5 h at 60° C. The reaction solution was combined with 5.7 g (23.3mmol) 3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one andstirred for 3 h at 100° C. The reaction mixture was evaporated downi.vac., the residue was combined with 200 mL EtOAc, the organic phasewas washed three times with 100 mL of 1 M KHSO₄ solution and 12 timeswith 50 mL 15% K₂CO₃ solution and dried over MgSO₄. After the desiccantand solvent had been eliminated the residue was further reacted withoutpurification.

Yield: 9.0 g (75% of theory)

ESI-MS: (M+H)⁺=549

R_(f)=0.64 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

14h) (R)-2-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 0.85 g (34.7 mmol) LiOH in 40 mL water was added to asolution of 9.0 g (16.4 mmol)(R)-2-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-1-methoxy-carbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 100 mL THF and the reaction mixture was stirred for 2 h at RT. TheTHF was eliminated i.vac., the residue was diluted with water, extractedtwice with 50 mL MTBE and the aqueous phase was acidified with 9 mL of 4M HCl. The precipitate was separated off, washed with water and dried.Further purification was carried out by trituration with 50 mL MTBE andfurther suction filtering of the product.

Yield: 7.5 g (86% of theory)

ESI-MS: (M+H)⁺=535

R_(f)=0.25 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

14i)(R)-1-(4-amino-3-methyl-5-trifluoromethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 100 mg (0.19 mmol)(R)-2-(4-amino-3-methyl-5-trifluoromethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate,67 mg (0.21 mmol) TBTU and 50 μL (0.36 mmol) triethylamine in 1 mL DMFwas stirred for 10 min at RT. Then 40 mg (0.24 mmol)1-(tetrahydropyran-4-yl)piperazine were added and the reaction mixturewas stirred for a further 20 h at RT. The reaction solution was purifiedby HPLC without any further working up. The fractions containing theproduct were combined and lyophilised.

Yield: 83 mg (65% of theory)

ESI-MS: (M+H)⁺=687

retention time (HPLC-MS):3.3 min (method A)

Example 15(S)-2-(4-amino-3-chloro-5-methyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

15a) ethyl 4-amino-3-bromo-5-chloro-benzoate

A solution of 20.0 g (81.9 mmol) ethyl 4-amino-3-bromo-benzoate in 150mL AcOH was heated to 40° C. Then 11.0 mL (133 mmol) sulphuryl chloridewere added dropwise in such a way that the internal temperature did notexceed 45° C. After the addition had ended the mixture was stirred for afurther 2 h at 45° C. The reaction mixture was evaporated down to about50 mL i.vac., poured onto ice water, the mixture was stirred for 10 min,the precipitate formed was filtered off and dried at 40° C.

Yield: 21.8 g (96% of theory)

R_(f)=0.62 (silica gel, PE/EtOAc 1:1)

15b) 4-amino-3-bromo-5-chloro-benzoic acid

A solution of 21.0 g (75.4 mmol) ethyl 4-amino-3-bromo-5-chloro-benzoatein 200 mL 4 M HCl and 100 mL EtOH was refluxed overnight. After coolingthe precipitate formed was suction filtered and dried.

Yield: 14.5 g (77% of theory)

ESI-MS: (M−H)⁻=248/250/252 (Br/Cl)

15c) (4-amino-3-bromo-5-chloro-phenyl)-methanol

10.3 g (63.8 mmol) CU were added to a solution of 14.5 g (57.9 mmol)4-amino-3-bromo-5-chloro-benzoic acid in 200 mL THF and the reactionmixture was stirred for 1 h at 40° C. The reaction mixture was left tocool to RT and then added to a solution of 7.67 g (203 mmol) sodiumborohydride in 200 mL water under a nitrogen atmosphere such that thetemperature did not exceed 30° C. After the addition had ended themixture was stirred for 2 h at RT, then diluted with 150 mL water,acidified with 100 mL 4 M HCl and stirred for a further hour at RT. Themixture was extracted twice with EtOAc and the combined organic phaseswere dried over Na₂SO₄. After the desiccant and solvent had beeneliminated the product was further reacted without purification.

Yield: 13.4 g (98% of theory)

ESI-MS: (M−H₂O+H)⁺=218/220/222 (Br/Cl)

R_(f)=0.44 (silica gel, DCM/MeOH/NH₃ 90:10:1)

15d) 4-amino-3-bromo-5-chloro-benzaldehyde

A solution of 13.4 g (56.7 mmol)(4-amino-3-bromo-5-chloro-phenyl)methanol in 300 mL DCM was combinedbatchwise with 78.0 g (897 mmol) manganese(IV) oxide while cooling withice and the resulting mixture was stirred for 4 h at RT. The reactionmixture was filtered and evaporated down i.vac. The product was furtherreacted without purification.

Yield: 12.9 g (97% of theory)

ESI-MS: (M+H)⁺=234/236/238 (Br/Cl)

R_(f)=0.92 (silica gel, DCM/MeOH/NH₃ 90:10:1)

15e) 1-methyl2-[1-(4-amino-3-bromo-5-chloro-phenyl)-meth-(Z)-ylidene]-succinate

Prepared analogously to Example 12b from 12.9 g (55.0 mmol)4-amino-3-bromo-5-chloro-benzaldehyde and 43.6 g (111 mmol) 1-methyl2-(triphenyl-λ⁵-phosphanyl idene)-succinate.

Yield: 12.5 g (65% of theory)

ESI-MS: (M−H)⁻=346/348/350 (Br/Cl)

R_(f)=0.63 (silica gel, EtOAc)

15f) 1-methyl (S)-2-(4-amino-3-bromo-5-chloro-benzyl)-succinate

Prepared analogously to Example 12c from 12.4 g (35.6 mmol) 1-methyl2-[1-(4-amino-3-bromo-5-chloro-phenyl)-meth-(Z)-ylidene]-succinate and450 mg(+1,2-bis((2R,5R)-2,5-diethylphospholano)benzene(cyclooctadiene)rhodium(I)tetrafluoroborate,the reaction mixture being hydrogenated for 20 h.

Yield: 11.3 g (91% of theory)

ESI-MS: (M−H)⁻=348/350/352 (Br/Cl)

retention time (HPLC-MS):7.1 min (method B)

15g) methyl(S)-2-(4-amino-3-bromo-5-chloro-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoate

5.4 g (22.0 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one were addedto a mixture of 7.0 g (20.0 mmol) 1-methyl(S)-2-(4-amino-3-bromo-5-chloro-benzyl)-succinate, 7.1 g (22.0 mmol)TBTU and 3.78 mL (22.0 mmol) ethyldiisopropylamine in 40 mL THF and thereaction mixture was shaken overnight at RT. The reaction solution wasevaporated down i.vac., the residue was combined with 15% K₂CO₃ solutionand treated in the ultrasound bath. The precipitate was suctionfiltered, washed with water, dried, taken up in a little DCM andpurified by chromatography (silica gel, gradient DCM to DCM/MeOH/NH₃70:30:3).

Yield: 8.4 g (73% of theory)

ESI-MS: (M+H)⁺=577/579/581(Br/Cl)

R_(f)=0.60 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

15h) methyl(S)-2-(4-amino-3-chloro-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoate

Under a nitrogen atmosphere 0.99 g (16.0 mmol) methylboric acid, 15.5 mL2 M Na₂CO₃ solution and 1.02 g (1.40 mmol) Pd(dppf)Cl₂ were added to asolution of 8.40 g (14.5 mmol) methyl(S)-2-(4-amino-3-bromo-5-chloro-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoatein 50 mL 1,4-dioxane and 3 mL MeOH and the reaction mixture was refluxedovernight. The reaction solution was filtered while hot and the filtratewas combined with EtOAc. The organic phase was washed several times withsemisaturated NaHCO₃ solution and dried over Na₂SO₄. It was filteredthrough activated charcoal and evaporated down i.vac. The residue wastaken up in a little DCM and purified by chromatography (silica gel,gradient DCM to DCM/MeOH/NH₃ 90:10:1). The fractions containing theproduct were combined, evaporated down i.vac., triturated with DIPE,suction filtered and dried.

Yield: 2.2 g (30% of theory)

ESI-MS: (M+H)⁺=513/515 (Cl)

retention time (HPLC-MS):4.0 min (method A)

15i)(S)-2-(4-amino-3-chloro-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid

A solution of 2.20 g (4.29 mmol) methyl(S)-2-(4-amino-3-chloro-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoatein 20 mL THF was combined with a solution of 156 mg (6.50 mmol) LiOH in5 mL water and stirred overnight at RT. The reaction mixture wasevaporated down i.vac., the residue was taken up in water, acidifiedwith 2 M HCl, the precipitate was suction filtered and dried in thevacuum drying cupboard at 40° C. This was taken up in a little DCM andpurified by chromatography (silica gel, gradient DCM/MeOH/NH₃ 90:10:1 toDCM/MeOH/NH₃ 70:30:3).

Yield: 1.3 g (61% of theory)

ESI-MS: (M+H)⁺=499/501 (Cl)

R_(f)=0.18 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

15k)(S)-2-(4-amino-3-chloro-5-methyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

Prepared analogously to Example 71 from 70 mg (0.14 mmol)(S)-2-(4-amino-3-chloro-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)piperidin-1-yl]-butanoicacid and 27 mg (0.16 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 40 mg (44% of theory)

ESI-MS: (M+H)⁺=651/653 (Cl)

retention time (HPLC-MS):2.6 min (method A)

Example 16(S)-2-(3-chloro-4-hydroxy-5-trifluoromethyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-Y^(1])-butane-1,4-dione

16a) 4-hydroxy-3-trifluoromethyl-benzoic acid

10.0 g (45.4 mmol) 4-methoxy-3-trifluoromethyl-benzoic acid and 75 gpyridinium-hydrochloride were mixed well and then heated to 180° C.under a nitrogen atmosphere for 5 h. The reaction mixture was pouredonto 1 L 10% citric acid solution and extracted with 50 mL EtOAc. Theorganic phase was washed with 1 L water and dried over Na₂SO₄. After thedesiccant and solvent had been eliminated the product was furtherreacted without purification.

Yield: 11.7 g

ESI-MS: (M−H)⁻=205

retention time (HPLC-MS):6.1 min (method B)

16b) 3-chloro-4-hydroxy-5-trifluoromethyl-benzoic acid

11.7 g of the crude product from Example 16a was dissolved in 40 mL AcOHat 40° C. At this temperature 5.15 mL (63 mmol) sulphuryl chloride wereadded dropwise and after the end of the addition the reaction mixturewas stirred for a further 2 h at this temperature. To complete thereaction another 2.5 mL sulphuryl chloride were added dropwise and thereaction mixture was heated to 60° C. for 4 h. The reaction solution waspoured onto 300 mL water, extracted with 200 mL EtOAc, the organic phasewas washed twice with water and dried over Na₂SO₄. After the desiccantand solvent had been eliminated the residue was stirred out with 80 mLPE, the precipitated substance was suction filtered, washed with 20 mLPE and dried.

Yield: 7.7 g (70% of theory over 2 steps)

ESI-MS: (M−H)⁻=239/241 (Cl)

retention time (HPLC-MS):6.5 min (method B)

16c) 2-chloro-4-hydroxymethyl-6-trifluoromethyl-phenol

5.76 g (36.0 mmol) CU were added to a solution of 7.70 g (32.0 mmol)3-chloro-4-hydroxy-5-trifluoromethyl-benzoic acid in 100 mL THF and thereaction mixture was stirred for 1 h at 40° C. After cooling to RT thissolution was carefully added to a solution of 3.78 g (100 mmol) sodiumborohydride in 40 mL water under a nitrogen atmosphere, such that thetemperature did not exceed 30° C. during the addition. After theaddition had ended the mixture was stirred for a further 2 h at RT,diluted with 200 mL water, acidified with 50 mL semiconcentrated HCl,stirred for 1 h, exhaustively extracted with EtOAc and the combinedorganic phases were dried over Na₂SO₄. After the desiccant and solventhad been eliminated the product was further reacted withoutpurification.

Yield: 5.9 g (81% of theory)

ESI-MS: (M−H)⁻=225/227 (Cl)

R_(f)=0.85 (silica gel, EtOAc)

16d) 3-chloro-4-hydroxy-5-trifluoromethyl-benzaldehyde

30.0 g (345 mmol) manganese(IV) oxide were added to a solution of 5.90 g(26.0 mmol) 2-chloro-4-hydroxymethyl-6-trifluoromethyl-phenol in 100 mLDCM and the reaction mixture was stirred for 2 h at RT. The precipitatewas filtered off, the filtrate was evaporated down i.vac. and furtherreacted without purification.

Yield: 3.0 g (51% of theory)

ESI-MS: (M−H)⁻=223/225 (Cl)

R_(f)=0.5 (silica gel, PE/EtOAc 1:1)

16e) 1-methyl2-[1-(3-chloro-4-hydroxy-5-trifluoromethyl-phenyl)-meth-(Z)-ylidene]-succinate

Prepared analogously to Example 12b from 3.0 g (13.4 mmol)3-chloro-4-hydroxy-5-trifluoromethyl-benzaldehyde and 10.5 g (26.7 mmol)1-methyl 2-(triphenyl-λ⁵-phosphanylidene)-succinate. The crude productobtained was purified by chromatography (silica gel, gradient PE/EtOAc1:1 to EtOAc).

Yield: 2.5 g (55% of theory)

ESI-MS: (M−H)⁻=337/339 (Cl)

R_(f)=0.75 (silica gel, EtOAc)

16f) 1-methyl(S)-2-(3-chloro-4-hydroxy-5-trifluoromethyl-benzyl)-succinate

Prepared analogously to Example 12c from 2.30 g (6.79 mmol) 1-methyl2-[1-(3-chloro-4-hydroxy-5-trifluoromethyl-phenyl)-meth-(Z)-ylidene]-succinateand 100 mg(−)-1,2-bis((2R,5R)-2,5-diethylphospholano)benzene(cyclooctadiene)rhodium(I)tetrafluoroborate.

Yield: 1.7 g (74% of theory)

ESI-MS: (M−H)⁻=339/341 (Cl)

retention time (HPLC-MS):7.1 min (method B)

16g) methyl(S)-2-(3-chloro-4-hydroxy-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoate

1.65 g (4.84 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one were addedto a mixture of 1.19 g (4.85 mmol) 1-methyl(S)-2-(3-chloro-4-hydroxy-5-trifluoromethyl-benzyl)-succinate, 1.56 g(4.85 mmol) TBTU, 0.73 mL (5.00 mmol) triethylamine, in 30 mL DMF andthe reaction mixture was stirred overnight at RT. The reaction solutionwas evaporated down i.vac., the residue was taken up in 200 mL EtOAc,the organic phase was washed with 10% citric acid and saturated Na₂CO₃solution and dried over Na₂SO₄. After the desiccant and solvent had beeneliminated the desired product was obtained.

Yield: 1.8 g (65% of theory)

ESI-MS: (M+H)⁺=568/570 (Cl)

retention time (HPLC-MS):8.1 min (method B)

16h)(S)-2-(3-chloro-4-hydroxy-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid

A solution of 115 mg (4.80 mmol) LiOH in 50 mL water was added to asolution of 1.80 g (3.17 mmol) methyl(S)-2-(3-chloro-4-hydroxy-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-Y^(1])-butanoatein 50 mL THF and the reaction mixture was stirred overnight at RT. Thereaction solution was freed from the THF i.vac., diluted with 150 mLwater, the aqueous phase was washed with 150 mL EtOAc, acidified withconcentrated HCl, extracted with 150 mL EtOAc, the organic phase wasseparated off and dried over Na₂SO₄. After the desiccant and solvent hadbeen eliminated the product was further reacted without purification.

Yield: 1.5 g (85% of theory)

ESI-MS: (M+H)⁺=554/556 (Cl)

retention time (HPLC-MS):8.2 min (method B)

16i)(S)-2-(3-chloro-4-hydroxy-5-trifluoromethyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

Prepared analogously to Example 71 from 70 mg (0.13 mmol)(S)-2-(3-chloro-4-hydroxy-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid and 22 mg (0.13 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 56 mg (63% of theory)

ESI-MS: (M+H)⁺=706/708 (Cl)

retention time (HPLC-MS):6.0 min (method B)

Example 17(S)-2-(4-hydroxy-3,5-dimethyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

17a) 4-benzyloxy-3,5-dimethyl-benzaldehyde

Under an argon atmosphere 60.8 g (440 mmol) K₂CO₃ and 52.3 mL (440 mmol)benzylbromide were added to a solution of 60.1 g (400 mmol)4-hydroxy-3,5-di-methyl-benzaldehyde in 600 mL acetone and the reactionmixture was heated to 50° C. for 2.5 h. The precipitate was filteredoff, washed with acetone and the filtrate was evaporated down i.vac. Theresidue was purified by chromatography (silica gel, Cyc/EtOAc 9:1).

Yield: 94.8 g (99% of theory)

ESI-MS: (M+H)⁺=241

R_(f)=0.45 (silica gel, Cyc/EtOAc 4:1)

17b) 1-methyl2-[1-(4-benzyloxy-3,5-dimethyl-phenyl)-meth-(Z)-ylidene]-succinate

Prepared analogously to Example 12b from 29.0 g (96.6 mmol)4-benzyloxy-3,5-dimethyl-benzaldehyde and 75.8 g (193 mmol) 1-methyl2-(triphenyl-λ⁵-phosphanylidene)-succinate.

Yield: 5.67 g (17% of theory)

ESI-MS: (M+H)⁺=355

17c) 1-methyl (S)-2-(4-benzyloxy-3,5-dimethyl-benzyl)-succinate

Under an argon atmosphere 100 mg(−)-1,2-bis((2R,5R)-2,5-diethylphospholano)benzene(cyclooctadiene)rhodium(I)tetrafluoroboratewere added to a solution of 5.67 g (16.0 mmol) 1-methyl2-[1-(4-benzyloxy-3,5-dimethyl-phenyl)-meth-(Z)-ylidene]-succinate in 40mL degassed MeOH and 5.0 mL triethylamine and the reaction mixture washydrogenated at 3447 hPa hydrogen pressure for 7 h. Then the reactionsolution was evaporated down i.vac., the residue was suspended in 80 mL15% K₂CO₃ solution, extracted with 80 mL EtOAc and the organic phase wasseparated off. The aqueous phase was acidified with 2 M HCl, extractedtwice with 40 mL EtOAc, the combined organic phases were washed withsaturated NaCl solution and dried over Na₂SO₄. After the desiccant andsolvent had been eliminated the desired product was obtained.

Yield: 1.69 g (30% of theory)

ESI-MS: (M−H)⁻=355

retention time (HPLC-MS):9.2 min (method B)

17d) methyl(S)-2-(4-benzyloxy-3,5-dimethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoate

A mixture of 1.69 g (4.74 mmol) 1-methyl(S)-2-(4-benzyloxy-3,5-dimethyl-benzyl)-succinate, 1.55 g (4.83 mmol)TBTU, 0.69 g (5.10 mmol) HOBt and 1.34 mL (7.71 mmol)ethyldiisopropylamine in 40 mL THF and 5 mL DMF was stirred for 16 h atRT. Then 1.16 g (4.74 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one was added tothe reaction mixture, which was then stirred for a further 2.5 h at RT.The reaction solution was combined with 40 mL EtOAc, the organic phasewas washed twice with 30 mL semisaturated NaHCO₃ and once with 40 mLsaturated NaCl solution washed and dried over Na₂SO₄. After thedesiccant and solvent had been eliminated the residue was purified bychromatography (silica gel, EtOAc/Cyc 3:1).

Yield: 2.43 g (88% of theory)

ESI-MS: (M+H)⁺=584

retention time (HPLC-MS):10.0 min (method B)

17e)(S)-2-(4-benzyloxy-3,5-dimethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid

A solution of 200 mg (8.35 mmol) LiOH in 40 mL water was added to asolution of 2.43 g (4.16 mmol) methyl(S)-2-(4-benzyloxy-3,5-dimethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoatein 80 mL THF and the reaction mixture was stirred for 1 h at RT. Themixture was evaporated down i.vac., the residue was taken up in 100 mLwater and acidified with 2 M HCl with stirring. The precipitate formedwas separated off and dried at 40° C.

Yield: 2.41 g (100% of theory)

ESI-MS: (M+H)⁺=570

retention time (HPLC-MS):9.0 min (method B)

17f)(S)-2-(4-hydroxy-3,5-dimethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid

A suspension of 2.41 g (4.23 mmol)(S)-2-(4-benzyloxy-3,5-dimethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid and 300 mg 10% Pd/C in 50 mL DCM was hydrogenated at RT and 3447hPa hydrogen pressure until the theoretical amount of hydrogen had beentaken up. The catalyst was filtered off and the filtrate was evaporateddown i.vac. The residue was triturated with DIPE, suction filtered anddried.

Yield: 1.88 g (93% of theory)

ESI-MS: (M+H)⁺=480

retention time (HPLC-MS):6.7 min (method B)

17g)(S)-2-(4-hydroxy-3,5-dimethyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

A mixture of 70 mg (0.15 mmol)(S)-2-(4-hydroxy-3,5-dimethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid, 52 mg (0.15 mmol) TBTU and 25 μL (0.18 mmol) triethylamine in 1 mLDMF was stirred for 1 h at RT. Then 25 mg (0.15 mmol)1-(tetrahydropyran-4-yl)-piperazine was added to the reaction mixture,which was then stirred for a further 16 h at RT. The reaction mixturewas purified by HPLC without any further working up; the fractionscontaining the product were combined and lyophilised.

Yield: 37 mg (40% of theory)

ESI-MS: (M+H)⁺=632

retention time (HPLC-MS):5.6 min (method B)

Example 18(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(6-hydroxy-2-oxo-1,4-dihydro-2H-quinazolin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

18a) methyl(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(6-hydroxy-2-oxo-1,4-dihydro-2H-quinazolin-3-yl)-piperidin-1-yl]-4-oxo-butanoate

A solution of 1.37 g (4.04 mmol) 1-methyl(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-succinate, 1.42 g(4.40 mmol) TBTU, 0.63 mL (4.50 mmol) triethylamine and 1.00 g (4.04mmol) 6-hydroxy-3-piperidin-4-yl-3,4-dihydro-1H-quinazolin-2-one in 10mL DMF was stirred for 3 h at RT. The reaction mixture was poured onto300 mL saturated NaHCO₃ solution, the precipitated substance was suctionfiltered, washed with 50 mL water and dried at 60° C. in the circulatingair dryer.

Yield: 2.30 g (100% of theory)

ESI-MS: (M+H)⁺=569/571 (Cl)

retention time (HPLC): 3.6 min (method A)

18b)(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(6-hydroxy-2-oxo-1,4-dihydro-2H-quinazolin-3-yl)-piperidin-1-yl]-4-oxo-butanoicacid

A solution of 144 mg (6.00 mmol) LiOH in 15 mL water was added to asolution of 2.30 g (4.04 mmol) methyl(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(6-hydroxy-2-oxo-1,4-dihydro-2H-quinazolin-3-yl)-piperidin-1-yl]-4-oxo-butanoatein 30 mL THF and the reaction mixture was stirred for 1 h at RT. Theorganic solvent was eliminated i.vac., the residue was diluted with 50mL water and acidified with 1 M HCl. The precipitate formed wasfiltered, washed with 10 mL water and dried at 50° C.

Yield: 2.20 g (98% of theory)

ESI-MS: (M+H)⁺=555/557 (Cl)

retention time (HPLC): 3.2 min (method A)

18c)(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(6-hydroxy-2-oxo-1,4-dihydro-2H-quinazolin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

Prepared analogously to Example 81 from 80.0 mg (0.14 mmol)(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(6-hydroxy-2-oxo-1,4-dihydro-2H-quinazolin-3-yl)-piperidin-1-yl]-4-oxo-butanoicacid and 24.5 mg (0.14 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 56 mg (55% of theory)

ESI-MS: (M+H)⁺=707/709 (Cl)

retention time (HPLC-MS):2.8 min (method A)

Example 19(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

19a) methyl(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidin-1-yl]-butanoate

A mixture of 3.00 g (8.83 mmol) 1-methyl(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-succinate, 3.05 g(9.50 mmol) TBTU and 1.7 mL (9.76 mmol) ethyldiisopropylamine in 100 mLDMF was stirred for 1 h at RT. Then 2.55 g (9.50 mmol)3-piperidin-4-yl-1,3-dihydro-imidazo[4,5-c]quinoline-2-one was added tothe reaction mixture, which was then stirred overnight at RT. Thereaction mixture was evaporated down i.vac., the residue was taken up inDCM, the organic phase was washed with 10% citric acid and 15% K₂CO₃solution and dried over Na₂SO₄. After filtration through activatedcharcoal and elimination of the solvent the product was further reactedwithout purification.

Yield: 5.20 g (100% of theory)

ESI-MS: (M+H)⁺=590/592 (Cl)

R_(f)=0.66 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

19b)(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidin-1-yl]-butanoicacid

A solution of 566 mg (13.22 mmol) LiOH*H₂O in 12 mL water was added to asolution of 5.20 g (8.81 mmol) methyl(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidin-1-Y^(1])-butanoatein 29 mL THF and the reaction mixture was stirred for 7 h at RT. Thiswas evaporated down i.vac., the residue was combined with 100 mL waterand acidified with 1 M HCl. The precipitate was suction filtered,dissolved again in EtOAc, extracted with 15% K₂CO₃ solution and theaqueous phase was acidified with 1 M HCl. The precipitate was suctionfiltered and dried.

Yield: 2.75 g (54% of theory)

ESI-MS: (M+H)⁺=576/578 (Cl)

R_(f)=0.09 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

19c)(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-[4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

Prepared analogously to Example 71 from 80 mg (0.14 mmol)(S)-2-(4-amino-3-chloro-5-trifluoromethyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidin-1-yl]-butanoicacid and 26.0 mg (0.15 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 38 mg (38% of theory)

ESI-MS: (M+H)⁺=728/730 (Cl)

retention time (HPLC-MS):2.3 min (method H)

Example 20(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidine-1-carboxylate

20a) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere 1.28 g (6.36 mmol) 4-nitrophenylchloroformate were added at RT to a solution of 0.78 g (6.36 mmol)4-dimethylaminopyridine in 100 mL pyridine and the mixture was stirredfor 1 h at RT. Then a solution of 2.00 g (6.36 mmol) methyl(R)-3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-hydroxy-propionate in 20 mLpyridine was added dropwise at RT and the reaction mixture was stirredfor 2 h after the end of the addition. Then 1.71 g (6.36 mmol)3-piperidin-4-yl-1,3-dihydro-imidazo[4,5-c]quinoline-2-one was added tothe reaction mixture, which was then heated to 100° C. for 4 h. Theprecipitate formed was filtered off, the filtrate was evaporated downi.vac., the residue was mixed with 200 mL EtOAc and 200 mL semisaturatedKHSO₄ solution, the product being obtained as a precipitate. This wassuction filtered and dried.

Yield: 2.50 g (65% of theory)

ESI-MS: (M+H)⁺=609

retention time (HPLC-MS):3.9 min (method A)

20b) (R)-1-carboxy-2-(4-benzyloxy-3,5-dimethyl-phenyl)-ethyl4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidine-1-carboxylate

A solution of 250 mg (10.42 mmol) LiOH in 10 mL water was added to asolution of 2.50 g (4.11 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidine-1-carboxylatein 20 mL THF and the reaction mixture was stirred for 2 h at RT. Theorganic solvent was removed i.vac., the aqueous residue was acidifiedwith 2 M HCL and combined with EtOAc/DCM (2:1). The precipitate formedwas filtered off and dried.

Yield: 1.84 g (75% of theory)

ESI-MS: (M+H)⁺=595

retention time (HPLC-MS):3.6 min (method A)

20c) (R)-1-carboxy-2-(4-hydroxy-3,5-dimethyl-phenyl)-ethyl4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidine-1-carboxylate

A suspension of 1.80 g (3.03 mmol)(R)-1-carboxy-2-(4-benzyloxy-3,5-dimethyl-phenyl)-ethyl4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidine-1-carboxylateand 300 mg 10% Pd/C in 30 mL THF and 30 mL MeOH was hydrogenated at 3000hPa hydrogen pressure and RT for 48 h. The catalyst was suctionfiltered, the filtrate was evaporated down and the residue was purifiedby HPLC; the fractions containing the product were combined andlyophilised.

Yield: 0.25 g (16% of theory)

ESI-MS: (M+H)⁺=505

retention time (HPLC-MS):2.6 min (method A)

20d)(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidine-1-carboxylate

A solution of 100 mg (0.15 mmol)(R)-1-carboxy-2-(4-hydroxy-3,5-dimethyl-phenyl)-ethyl4-(2-oxo-1,2-dihydro-imidazo[4,5-c]quinolin-3-yl)-piperidine-1-carboxylate,53 mg (0.16 mmol) TBTU and 26 μL (0.19 mmol) triethylamine in 1 mL DMFwas stirred for 15 min at RT. Then 28 mg (0.16 mmol)1-(tetrahydropyran-4-yl)-piperazine was added and the reaction solutionwas stirred for 2 h. The reaction solution was purified by HPLC withoutany further working up; the fractions containing the product werecombined and lyophilised.

Yield: 42 mg (43% of theory)

ESI-MS: (M+H)⁺=657

retention time (HPLC-MS):2.9 min (method C)

Example 21(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate

21a) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere 0.91 g (4.38 mmol) 4-nitrophenylchloroformate in 10 mL THF were metered into 20 mL pyridine within 10min at a bath temperature of 60° C., the mixture was stirred for 10 min,then 1.38 g (4.38 mmol) methyl(R)-3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-hydroxy-propionate in 40 mLpyridine were added and the reaction mixture was stirred for 2 h at 60°C. The reaction solution was combined with 1.00 g (4.38 mmol)3-piperidin-4-yl-1H-quinoline-2-one and stirred for 4 h at 100° C. Thereaction mixture was evaporated down i.vac. and the residue was purifiedby HPLC. The fractions containing the product were combined, evaporateddown i.vac., the residue was made alkaline with 15% K₂CO₃ solution, theprecipitate was suction filtered, washed with 20 mL water and dried at50° C.

Yield: 0.62 g (25% of theory)

ESI-MS: (M+H)⁺=569

retention time (HPLC-MS):5.1 min (method A)

21b) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate

A solution of 38 mg (1.60 mmol) LiOH in 50 mL water was added to asolution of 600 mg (1.06 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate in 30 mLTHF and the reaction mixture was stirred for 1 h at RT. The organicsolvent was eliminated i.vac., the aqueous residue was diluted with 50mL water and acidified with 1 M HCL. The precipitate formed was filteredoff, washed with 10 mL water and dried at 50° C.

Yield: 600 mg (100% of theory)

ESI-MS: (M+H)⁺=555

retention time (HPLC-MS):4.3 min (method A)

21c) (R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate

A suspension of 600 mg (1.08 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate and 250 mgof 10% Pd/C in 50 mL isopropanol was hydrogenated at 50° C. and 3447 hPahydrogen pressure for 2 h. The catalyst was suction filtered, thefiltrate was evaporated down i.vac., the residue was triturated with 50mL diethyl ether, suction filtered, washed with 20 mL diethyl ether anddried at 50° C.

Yield: 430 mg (86% of theory)

ESI-MS: (M+H)⁺=465

retention time (HPLC-MS):3.4 min (method A)

21d)(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 81 from 80.0 mg (0.17 mmol)(R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate and 29.2 mg(0.17 mmol) 1-(tetrahydropyran-4-yl)-piperazine, the reaction mixturebeing stirred overnight at RT.

Yield: 52 mg (49% of theory)

ESI-MS: (M+H)⁺=617

retention time (HPLC-MS):3.0 min (method A)

Example 22(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(7-methoxy-2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

22a) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(7-methoxy-2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carbon/late

Prepared analogously to Example 21a from 1.41 g (3.63 mmol) methyl(R)-3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-hydroxy-propionate, 0.75 g(3.63 mmol) 4-nitrophenyl chloroformate and 1.00 g (3.63 mmol)7-methoxy-3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one.

Yield: 0.65 g (29% of theory)

ESI-MS: (M+H)⁺=616

retention time (HPLC-MS):5.1 min (method A)

22b) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(7-methoxy-2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 21b from 0.65 g (1.06 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-methoxycarbonyl-ethyl4-(7-methoxy-2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 38.4 mg (1.60 mmol) LiOH.

Yield: 0.64 g (100% of theory)

ESI-MS: (M+H)⁺=602

retention time (HPLC-MS):4.5 min (method A)

22c) (R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(7-methoxy-2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 21c from 0.64 g (1.06 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(7-methoxy-2-oxo-1,2,4,5-tetra-hydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 100 mg 10% Pd/C.

Yield: 0.50 g (92% of theory)

ESI-MS: (M+H)⁺=512

retention time (HPLC-MS):3.5 min (method A)

22d)(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(7-methoxy-2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 81 from 80.0 mg (0.17 mmol)(R)-2-(4-hydroxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2-dihydro-quinolin-3-yl)-piperidine-1-carboxylate and 29.0 mg(0.17 mmol) 1-(tetrahydropyran-4-yl)-piperazine, the reaction mixturebeing stirred overnight at RT.

Yield: 78 mg (75% of theory)

ESI-MS: (M+H)⁺=664

retention time (HPLC-MS):2.9 min (method A)

Example 23(S)-2-(4-hydroxy-3-methoxy-5-methyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

23a) 1-methyl2-[1-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-meth-(Z)-ylidene]-succinate

Prepared analogously to Example 12b from 9.80 g (38.2 mmol)4-benzyloxy-3-methoxy-5-methyl-benzaldehyde (Example 9c) and 45.0 g (115mmol) 1-methyl 2-(triphenyl-λ⁵-phosphanylidene)-succinate.

Yield: 13.6 g (96% of theory)

retention time (HPLC-MS):12.5 min (method D)

23b) 1-methyl (S)-2-(4-benzyloxy-3-methoxy-5-methyl-benzyl)-succinate

Prepared analogously to Example 12c from 6.15 g (16.6 mmol) 1-methyl2-[1-(4-benzyloxy-3-methoxy-5-methyl-phenyl)-meth-(Z)-ylidene]-succinate,using 82 mg (0.17 mmol) bis-(1,5-cyclooctadiene)dirhodium(I) dichlorideas catalyst and 92 mg (0.17 mmol) tert-butyl(2S,4S)-4-diphenylphosphanyl-2-[(diphenylphosphanyl)-methyl]-pyrrolidine-1-carboxylateas ligand.

Yield: 5.8 g (94% of theory)

ESI-MS: (M+H)⁺=373

retention time (HPLC-MS):12.1 min (method D)

23c) methyl(S)-2-(4-benzyloxy-3-methoxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoate

Prepared analogously to Example 12d from 5.80 g (15.6 mmol) 1-methyl(S)-2-(4-benzyloxy-3-methoxy-5-methyl-benzyl)-succinate and 4.20 g(17.13 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one, using 3.43g (17.9 mmol) (3-dimethylamino-propyl)-ethyl-carbodiimide and 2.38 g(19.5 mmol) 4-dimethylamino-pyridine for the coupling and 130 mLacetonitrile and 50 mL THF as solvent.

Yield: 7.8 g (84% of theory)

ESI-MS: (M+H)⁺=600

retention time (HPLC-MS):13.2 min (method D)

23d)(S)-2-(4-benzyloxy-3-methoxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid

Prepared analogously to Example 7g from 7.83 g (13.1 mmol) methyl(S)-2-(4-benzyloxy-3-methoxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoateand 1.25 g (52.2 mmol) LiOH.

Yield: 7.6 g (99% of theory)

ESI-MS: (M+H)⁺=586

retention time (HPLC-MS):11.7 min (method D)

23e)(S)-2-(4-hydroxy-3-methoxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid

A suspension of 7.60 g (12.98 mmol)(S)-2-(4-benzyloxy-3-methoxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid and 0.76 g 10% Pd/C in 2 mL triethylamine and 150 mL MeOH washydrogenated at RT and 2620 hPa hydrogen pressure for 16 h. To completethe reaction a further 0.38 g 10% Pd/C were added and the mixture wasagain hydrogenated for 3 h at RT. The catalyst was filtered off throughCelite and the filtrate was concentrated by evaporation i.vac. Theproduct was further reacted without purification.

Yield: 7.2 g (81% of theory)

ESI-MS: (M+H)⁺=496

retention time (HPLC-MS):7.7 min (method D)

23f)(S)-2-(4-hydroxy-3-methoxy-5-methyl-benzyl)-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-1-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-butane-1,4-dione

Prepared analogously to Example 71 from 100 mg (0.20 mmol)(S)-2-(4-hydroxy-3-methoxy-5-methyl-benzyl)-4-oxo-4-[4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidin-1-yl]-butanoicacid and 58.9 mg (0.24 mmol) 1-(tetrahydropyran-4-yl)-piperazine (usedas the bis-hydrochloride salt), using 84.4 mg (0.22 mmol) HATU as thecoupling reagent and 111 μL (0.65 mmol) ethyldiisopropylamine as thebase.

Yield: 91 mg (56% of theory)

ESI-MS: (M+H)⁺=648

R_(f)=0.70 (silica gel, DCM/MeOH/NH₃ 80:20:2)

Example 24(R)-1-(3,5-dibromo-4-hydroxy-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

24a) (Z,E)-3-(4-acetoxy-3,5-dibromo-phenyl)-2-acetylamino-acrylic acid

Prepared analogously to Example 9d from 30.0 g (107 mmol)3,5-dibromo-4-hydroxy-benzaldehyde and 18.8 g (161 mmol)N-acetylglycine. After the reaction mixture was cooled, the productprecipitated out and was filtered, washed with water and dried.

Yield: 35.7 g (79% of theory)

ESI-MS: (M+H)⁺=420/422/424 (2 Br)

R_(f)=0.20 (silica gel, DCM/MeOH/AcOH 90:10:1)

24b) 3-(3,5-dibromo-4-hydroxyphenyl)-2-oxo-propionic acid

Prepared analogously to Example 7c from 35.7 g (84.8 mmol)(Z,E)-3-(4-acetoxy-3,5-dibromo-phenyl)-2-acetylamino-acrylic acid and325 mL of 4 M HCl, using 290 mL NMP as solvent.

Yield: 20.5 g (72% of theory)

ESI-MS: (M−H)⁻=335/337/339 (2 Br)

R_(f)=0.35 (silica gel, DCM/MeOH/AcOH 80:20:2)

24c) (R)-3-(3,5-dibromo-4-hydroxy-phenyl)-2-hydroxy-propionic acid

Prepared analogously to Example 7d from 14.5 g (42.9 mmol)3-(3,5-dibromo-4-hydroxy-phenyl)-2-oxo-propionic acid and 30.9 g (96.3mmol) (1R)-B-chlorodiisopinocampheylborane.

Yield: 12.7 g (87% of theory)

ESI-MS: (M−H)⁻=337/339/341 (2 Br)

R_(f)=0.4 (silica gel, DCM/MeOH/AcOH 80:20:2)

retention time (HPLC-MS):6.4 min (method G)

24d) methyl (R)-3-(3,5-dibromo-4-hydroxy-phenyl)-2-hydroxy-propionate

Prepared analogously to Example 7e from 14.0 g (34.8 mmol)(R)-3-(3,5-dibromo-4-hydroxy-phenyl)-2-hydroxy-propionic acid, usingmethanolic HCl (6 M).

Yield: 7.0 g (57% of theory)

ESI-MS: (M−H)⁻=351/353/355 (2 Br)

retention time (HPLC-MS):9.8 min (method G)

24e) methyl(R)-3-[3,5-dibromo-4-(2-trimethylsilanyl-ethoxymethoxy)-phenyl]-2-hydroxy-propionate

Under a nitrogen atmosphere 11.1 g (76.6 mmol) 40% KF/Al₂O₃ were addedto a solution of 6.78 g (19.2 mmol) methyl(R)-3-(3,5-dibromo-4-hydroxy-phenyl)-2-hydroxy-propionate in 100 mLacetonitrile and the resulting suspension was stirred for a few min atRT. Then a solution of 4.07 mL (23.0 mmol)(2-chloromethoxyethyl)-trimethyl-silane in 20 mL acetonitrile was addedand the reaction mixture was stirred for 20 h at RT. The mixture wasfiltered through Celite, the solvent was evaporated down i.vac. and theresidue was purified by chromatography (silica gel, n-hexane/EtOAc 7:3).

Yield: 5.49 g (59% of theory)

R_(f)=0.45 (silica gel, n-hexane/EtOAc 1:1)

24f)(R)-2-[3,5-dibromo-4-(2-trimethylsilanyl-ethoxymethoxy)-phenyl]-1-methoxy-carbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 7f from 4.63 g (9.56 mmol) methyl(R)-3-[3,5-dibromo-4-(2-trimethylsilanyl-ethoxymethoxy)-phenyl]-2-hydroxy-propionateand 2.35 g (9.56 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one, using 1.23g (10.04 mmol) 4-dimethylaminopyridine as base and acetonitrile assolvent.

Yield: 4.35 g (69% of theory)

ESI-MS: (M+H)⁺=754/756/758 (2 Br)

retention time (HPLC): 29.2 min (method G)

24g) (R)-2-(3,5-dibromo-4-hydroxy-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Under a nitrogen atmosphere 5.46 mL methanolic H₂SO₄ (0.5 M) was addedto a solution of 4.30 g (5.69 mmol)(R)-2-[3,5-d]bromo-4-(2-trimethylsilanyl-ethoxymethoxy)-phenyl-1-methoxy-carbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylatein 40 mL THF and 40 mL MeOH and the reaction solution was stirred for 6h at RT. The reaction mixture was evaporated down i.vac. and the residuewas further reacted without purification.

Yield: quantitative

ESI-MS: (M+H)⁺=624/626/628 (2 Br)

retention time (HPLC): 17.3 min (method G)

24h) (R)-1-carboxy-2-(3,5-dibromo-4-hydroxy-phenyl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 0.51 g (21.3 mmol) LiOH was added to a solution of(R)-2-(3,5-dibromo-4-hydroxy-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate(crude product from Example 24g) in 80 mL THF and the reaction mixturewas stirred for 3 h at RT. The THF was eliminated i.vac., the aqueousphase was washed with EtOAc, acidified with 10% HCl and the aqueousphase was extracted exhaustively with EtOAc. The combined organic phaseswere evaporated down i.vac., suspended in diethyl ether, filtered, theresidue was dried and then purified by chromatography (silica gel,DCM/MeOH/AcOH 90:10:1).

Yield: 3.5 g (100% of theory)

ESI-MS: (M+H)⁺=610/612/614 (2 Br)

retention time (HPLC): 14.1 min (method G)

24i)(R)-1-(3,5-dibromo-4-hydroxy-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 81 from 100 mg (0.16 mmol)(R)-1-carboxy-2-(3,5-dibromo-4-hydroxy-phenyl)-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 30.6 mg (0.18 mmol) 1-(tetrahydropyran-4-yl)-piperazine, thereaction mixture being stirred overnight at RT.

Yield: 72 mg (58% of theory)

ESI-MS: (M+H)⁺=762/764/766 (2 Br)

retention time (HPLC-MS):3.0 min (method A)

Example 25(R)-1-(3-bromo-4-hydroxy-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

25a) (Z,E)-3-(4-acetoxy-3-bromo-phenyl)-2-acetylamino-acrylic acid

Prepared analogously to Example 9d from 75.0 g (366 mmol)3-bromo-4-hydroxy-benzaldehyde and 64.2 g (548 mmol) N-acetylglycine.After the reaction mixture had cooled the product precipitated out andwas then filtered, washed with water and dried.

Yield: 69.8 g (56% of theory)

retention time (HPLC): 7.6 min (method G)

25b) 3-(3-bromo-4-hydroxy-phenyl)-2-oxo-propionic acid

Prepared analogously to Example 24b from 69.7 g (204 mmol)(Z,E)-3-(4-acetoxy-3-bromo-phenyl)-2-acetylamino-acrylic acid and 750 mL4 M HCl.

Yield: 45.8 g (87% of theory)

retention time (HPLC): 7.8 min (method G)

25c) (R)-3-(3-bromo-4-hydroxy-phenyl)-2-hydroxy-propionic acid

Prepared analogously to Example 7d from 45.0 g (174 mmol)3-(3-bromo-4-hydroxy-phenyl)-2-oxo-propionic acid and 114.2 g (356 mmol)(1R)-B-chlorodiisopinocampheylborane.

Yield: 53.7 g (89% of theory)

retention time (HPLC): 4.0 min (method G)

25d) methyl (R)-3-(3-bromo-4-hydroxy-phenyl)-2-hydroxy-propionate

2.5 mL concentrated sulphuric acid were added to a solution of 53.6 g(154 mmol) (R)-3-(3-bromo-4-hydroxy-phenyl)-2-hydroxy-propionic acid in250 mL MeOH and the reaction mixture was stirred for 4 h at RT. Themixture was evaporated down i.vac., the residue was taken up in 250 mLEtOAc, the organic phase was washed twice with 100 mL saturated NaHCO₃solution and saturated NaCl solution and dried over Na₂SO₄. After thedesiccant and solvent had been eliminated the residue was furtherreacted without purification.

Yield: quantitative

retention time (HPLC): 6.8 min (method G)

25e) methyl(R)-3-[3-bromo-4-(2-trimethylsilanyl-ethoxymethoxy)-phenyl]-2-hydroxy-propionate

6.7 mL (39.1 mmol) ethyldiisopropylamine were added to a solution of10.2 g (34.6 mmol) methyl(R)-3-(3-bromo-4-hydroxy-phenyl)-2-hydroxy-propionate in 100 mL DCM andthe reaction mixture was cooled in the ice bath. Then a solution of 7.9mL (44.6 mmol) (2-chloromethoxy-ethyl)-trimethyl-silane in 20 mL DCM wasadded. The reaction mixture was stirred for 3 h at RT and then combinedwith another 0.67 mL ethyldiisopropylamine and 0.8 mL (4.5 mmol)(2-chloromethoxy-ethyl)-trimethyl-silane to complete the reaction andstirred for 1.5 h at RT. The reaction mixture was washed with 5% Na₂CO₃solution and saturated NaCl solution and dried over Na₂SO₄. After thedesiccant and solvent had been eliminated the residue was purified bychromatography (silica gel, Cyc/EtOAc 75:25).

Yield: 9.6 g (68% of theory)

retention time (HPLC): 15.1 min (method D)

25f)(R)-2-[3-bromo-4-(2-trimethylsilanyl-ethoxymethoxy)-phenyl]-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 24f from 4.55 g (11.2 mmol) methyl(R)-3-[3-bromo-4-(2-trimethylsilanyl-ethoxymethoxy)-phenyl]-2-hydroxy-propionateand 2.75 g (11.2 mmol)3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one.

Yield: 5.46 g (72% of theory)

retention time (HPLC): 16.5 min (method D)

25g) (R)-2-(3-bromo-4-hydroxy-phenyl)-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 24g from 5.40 g (7.98 mmol)(R)-2-[3-bromo-4-(2-trimethylsilanyl-ethoxy-methoxy)-phenyl]-1-methoxycarbonyl-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 7.7 mL (4.2 mmol) methanolic sulphuric acid (0.5 M). The crudeproduct (5.44 g) was further reacted without purification.

Yield: quantitative

retention time (HPLC): 9.9 min (method D)

25h) (R)-2-(3-bromo-4-hydroxy-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

A solution of 0.84 g (34.1 mmol) LiOH in 20 mL water was added to asolution of 5.44 g of the crude product from Example 25g in 80 mL THFand the reaction mixture was stirred for 1 h at RT. The organic solventwas eliminated i.vac., the aqueous phase was washed with EtOAc,acidified with 10% HCl and exhaustively extracted with EtOAc. Thecombined organic phases were washed with saturated NaCl solution anddried over Na₂SO₄. After the desiccant and solvent had been eliminatedthe residue was triturated with 90 mL diethyl ether, filtered, the solidwas washed with diethyl ether and dried at 45° C.

Yield: 4.10 g (89% of theory)

retention time (HPLC): 8.2 min (method D)

25i)(R)-1-(3-bromo-4-hydroxy-benzyl)-2-oxo-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylate

Prepared analogously to Example 71 from 100 mg (0.19 mmol)(R)-2-(3-bromo-4-hydroxy-phenyl)-1-carboxy-ethyl4-(2-oxo-1,2,4,5-tetrahydro-1,3-benzodiazepin-3-yl)-piperidine-1-carboxylateand 35.1 mg (0.21 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 58 mg (45% of theory)

ESI-MS: (M+H)⁺=684/686 (Br)

retention time (HPLC-MS):2.4 min (method A)

Example 26(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-2-oxo-ethyl1′,2′-dihydro-2′-oxospiro-4H-3′,1-benzoxazine-4,4′-piperidine-1-carboxylate

26a) (R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl1′,2′-dihydro-2′-oxospiro-4H-3′,1-benzoxazine-4,4′-piperidine-1-carboxylate

Prepared analogously to Example 2g from 2.00 g (6.36 mmol) methyl(R)-3-(4-benzyloxy-3,5-dimethyl-phenyl)-2-hydroxy-propionate and 1.63 g(6.40 mmol) 3-piperidin-4-yl-1,3,4,5-tetrahydro-1,3-benzodiazepin-2-one.The hydrolysis of the methyl ester was carried out with 310 mg (12.95mmol) LiOH.

Yield: 1.00 g (29% of theory)

ESI-MS: (M+NH₄)⁺=562

R_(f)=0.12 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

26b)(R)-1-(4-benzyloxy-3,5-dimethyl-benzyl)-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-2-oxo-ethyl1′,2′-dihydro-2′-oxospiro-4H-3′,1-benzoxazine-4,4′-piperidine-1-carboxylate

Prepared analogously to Example 81 from 200 mg (0.37 mmol)(R)-2-(4-benzyloxy-3,5-dimethyl-phenyl)-1-carboxy-ethyl1′,2′-dihydro-2′-oxospiro-4H-3′,1-benzoxazine-4,4′-piperidine-1-carboxylateand 70.0 mg (0.41 mmol) 1-(tetrahydropyran-4-yl)-piperazine.

Yield: 130 mg (51% of theory)

ESI-MS: (M+H)⁺=697

R_(f)=0.42 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

26c)(R)-1-(4-hydroxy-3,5-dimethyl-benzyl)-2-[4-(tetrahydropyran-4-yl)-piperazin-1-yl]-2-oxo-ethyl1′,2′-dihydro-2′-oxospiro-4H-3′,1-benzoxazine-4,4′-piperidine-1-carboxylate

A suspension of 120 mg (0.17 mmol)(R)-1-(4-benzyloxy-3,5-dimethyl-benzyl)-2-[4-(tetrahydropyran-4-yl)piperazin-1-yl]-2-oxo-ethyl1′,2′-dihydro-2′-oxospiro-4H-3′,1-benzoxazine-4,4′-piperidine-1-carboxylateand 20 mg 10% Pd/C in 20 mL MeOH was hydrogenated at RT and 3000 hPahydrogen pressure for 7.5 h. The catalyst was suction filtered and thefiltrate was concentrated by evaporation. The residue was trituratedwith DIPE, suction filtered and dried at 50° C.

Yield: 95 mg (91% of theory)

ESI-MS: (M+H)⁺=607

R_(f)=0.41 (silica gel, DCM/Cyc/MeOH/NH₃ 70:15:15:2)

The following Examples describe the preparation of pharmaceuticalformulations which contain as active substance any desired compound ofgeneral formula I:

Example I Capsules for Powder Inhalation Containing 1 mg of ActiveIngredient Composition:

1 capsule for powder inhalation contains:

active ingredient  1.0 mg lactose 20.0 mg hard gelatine capsules 50.0 mg71.0 mg

Method of Preparation:

The active ingredient is ground to the particle size required forinhaled substances. The ground active ingredient is homogeneously mixedwith the lactose. The mixture is transferred into hard gelatinecapsules.

Example II Inhalable Solution for Respimat® Containing 1 mg of ActiveIngredient Composition:

1 spray contains:

active ingredient 1.0 mg benzalkonium chloride 0.002 mg disodium edetate0.0075 mg purified water ad 15.0 μl

Method of Preparation:

The active ingredient and benzalkonium chloride are dissolved in waterand transferred into Respimat® cartridges.

Example III Inhalable Solution for Nebulisers Containing 1 mg of ActiveIngredient Composition:

1 vial contains:

active ingredient 0.1 g sodium chloride 0.18 g benzalkonium chloride0.002 g purified water ad 20.0 ml

Method of Preparation:

The active ingredient, sodium chloride and benzalkonium chloride aredissolved in water.

Example IV Propellant Gas-Operated Metering Aerosol Containing 1 mg ofActive Ingredient Composition:

1 puff contains:

active ingredient  1.0 mg lecithin 0.1% propellant gas ad 50.0 μl

Method of Preparation:

The micronised active ingredient is homogeneously suspended in themixture of lecithin and propellant gas. The suspension is transferredinto a pressurised container with a metering valve.

Example V Nasal Spray Containing 1 mg of Active Ingredient Composition:

active ingredient 1.0 mg sodium chloride 0.9 mg benzalkonium chloride0.025 mg disodium edetate 0.05 mg purified water ad 0.1 ml

Method of Preparation:

The active ingredient and the excipients are dissolved in water andtransferred into a suitable container.

Example VI Injectable Solution Containing 5 mg of Active Substance per 5ml Composition:

active substance 5 mg glucose 250 mg human serum albumin 10 mgglycofurol 250 mg water for injections ad 5 ml

Preparation:

Glycofurol and glucose are dissolved in water for injections (WfI);human serum albumin is added; active ingredient is dissolved withheating; made up to specified volume with WfI; transferred into ampoulesunder nitrogen gas.

Example VII Injectable Solution Containing 100 mg of Active Substanceper 20 ml Composition:

active substance 100 mg monopotassium dihydrogen phosphate = KH₂PO₄ 12mg disodium hydrogen phosphate = Na₂HPO₄*2H₂O 2 mg sodium chloride 180mg human serum albumin 50 mg Polysorbate 80 20 mg water for injectionsad 20 ml

Preparation:

Polysorbate 80, sodium chloride, monopotassium dihydrogen phosphate anddisodium hydrogen phosphate are dissolved in water for injections (WfI);human serum albumin is added; active ingredient is dissolved withheating; made up to specified volume with WfI; transferred intoampoules.

Example VIII Lyophilisate Containing 10 mg of Active SubstanceComposition:

Active substance  10 mg Mannitol 300 mg human serum albumin  20 mg waterfor injections ad  2 ml

Preparation:

Mannitol is dissolved in water for injections (WfI); human serum albuminis added; active ingredient is dissolved with heating; made up tospecified volume with WfI; transferred into vials; freeze-dried.

Solvent for Lyophilisate:

Polysorbate 80 = Tween 80  20 mg mannitol 200 mg water for injections ad 10 ml

Preparation:

Polysorbate 80 and mannitol are dissolved in water for injections (WfI);transferred into ampoules.

Example IX Tablets Containing 20 mg of Active Substance Composition:

active substance 20 mg lactose 120 mg  corn starch 40 mg magnesiumstearate  2 mg Povidone K 25 18 mg

Preparation:

Active substance, lactose and corn starch are homogeneously mixed;granulated with an aqueous solution of Povidone; mixed with magnesiumstearate; compressed in a tablet press; weight of tablet 200 mg.

Example X Capsules Containing 20 mg Active Substance Composition:

active substance 20 mg corn starch 80 mg highly dispersed silica  5 mgmagnesium stearate 2.5 mg 

Preparation:

Active substance, corn starch and silica are homogeneously mixed; mixedwith magnesium stearate; the mixture is packed into size for 3 hardgelatine capsules in a capsule filling machine.

Example XI Suppositories Containing 50 mg of Active SubstanceComposition:

active substance  50 mg hard fat (Adeps solidus) q.s. ad 1700 mg

Preparation:

Hard fat is melted at about 38° C.; ground active substance ishomogeneously dispersed in the molten hard fat; after cooling to about35° C. it is poured into chilled moulds.

Example XII Injectable Solution Containing 10 mg of Active Substance per1 ml Composition:

active substance 10 mg mannitol 50 mg human serum albumin 10 mg waterfor injections ad  1 ml

Preparation:

Mannitol is dissolved in water for injections (WfI); human serum albuminis added; active ingredient is dissolved with heating; made up tospecified volume with WfI; transferred into ampoules under nitrogen gas.

1. A compound of the formula

wherein X denotes CH₂, NH, C₁₋₃-alkyl-N, O or S, R¹ denotes a moietyselected from the group consisting of

wherein R^(1.1) denotes H, halogen, HO, F₃C or C₁₋₆-alkyl-O, R² denotesa group of the formulae II

wherein R^(2.1) denotes H, halogen, C₁₋₃-alkyl-O, C₁₋₃-alkyl or F₃C,R^(2.2) denotes H, H₂N, HO, H₃C—O, H—C(O)—O or C₁₋₃-alkyl-C(O)-O,R^(2.3) denotes H, halogen, C₁₋₃-alkyl or F₃C, or R² denotes a moietyselected from the group consisting of

wherein R^(2.4) denotes H or H₃C, R³ denotes a group of the formulae III

wherein R^(3.1) denotes H, C₁₋₃-alkyl or R^(3.1.1)—(O)C, R^(3.1.1)denotes HO or C₁₋₆-alkyl-O, R^(3.2) denotes H or C₁₋₃-alkyl and R^(3.3)denotes a free pair of electrons or the oxygen atom, R⁴ denotes a 4- to7-membered oxycycloalkyl group optionally substituted by R^(4.1) andR^(4.1) denotes NC, HO, C₁₋₃-alkyl or C₁₋₃-alkyl-O, or a tautomer orsalt thereof.
 2. A compound of the formula I according to claim 1,wherein X denotes CH₂, NH or O, R¹ denotes a moiety selected from thegroup consisting of

wherein R^(1.1) denotes H, Cl, Br, HO, F₃C or H₃C—O, R² denotes a groupof the formulae II

wherein R^(2.1) denotes H, Cl, Br, H₃C—O, H₃C, F₃C or H₃C—H₂C, R^(2.2)denotes H₂N, HO, H₃C—O, H—C(O)—O or H₃C—C(O)—O, R^(2.3) denotes H, Cl,Br, H₃C or F₃C, or R² denotes a moiety selected from the groupconsisting of

wherein R^(2.4) denotes H or H₃C, R³ denotes a group of the formulae III

wherein R^(3.1) denotes H or H₃C, R^(3.2) denotes H or H₃C and R^(3.3)denotes a free pair of electrons or the oxygen atom, R⁴ denotes a 4- to7-membered oxycycloalkyl group optionally substituted by R^(4.1) andR^(4.1) denotes HO or C₁₋₃-alkyl, or a tautomer or salt thereof.
 3. Acompound of the formula I according to claim 1, wherein X denotes CH₂,NH or O, R¹ denotes a moiety selected from the group consisting of

wherein R^(1.1) denotes H or H₃C—O, R² denotes a moiety selected fromthe group consisting of

wherein R^(2.1) denotes H₃C or F₃C, R^(2.2) denotes H₂N or HO, R^(2.4)denotes H or H₃C, R³ denotes a moiety selected from the group consistingof

and R⁴ denotes a moiety selected from the group consisting of

or a tautomer or salt thereof.
 4. A compound of the formula I accordingto claim 1, wherein R¹ denotes a moiety selected from the groupconsisting of

R² denotes a moiety selected from the group consisting of

R³ denotes a moiety selected from the group consisting of

R⁴ denotes a moiety selected from the group consisting of

or a tautomer or salt thereof.
 5. A compound of the formula I accordingto claim 1, wherein R¹ denotes a moiety selected from the groupconsisting of

R² denotes a moiety selected from the group consisting of

R³—R⁴ together represent a moiety selected from the group consisting of

or a tautomer or salt thereof.
 6. A compound selected from the groupconsisting of: No. Structure (1)

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or a tautomer or salt thereof.
 7. A physiologically acceptable salt of acompound according to claim 1, 2, 3, 4, 5 or
 6. 8. A pharmaceuticalcomposition containing a compound according to claim 1, 2, 3, 4, 5 or 6or a physiologically acceptable salt thereof together with one or moreinert carriers and/or diluents.
 9. A method for the treatment ofmigraine, cluster headache and tension headache, which comprises theadministration to a person suffering from the same a therapeutic amountof a compound according to claim 1, 2, 3, 4, 5 or 6 or a physiologicallyacceptable salt thereof together.
 10. A method for the treatment ofnon-insulin-dependent diabetes mellitus (NIDDM), which comprises theadministration to a person suffering from the same a therapeutic amountof a compound according to claim 1, 2, 3, 4, 5 or 6 or a physiologicallyacceptable salt thereof together.
 11. A method for the treatment ofcardiovascular diseases, morphine tolerance, diarrhoea caused byclostridium toxin, skin diseases, particularly thermal andradiation-induced skin damage including sunburn, lichen, prurigo,pruriginous toxidermies and severe itching, inflammatory diseases, e.g.inflammatory diseases of the joints such as osteoarthritis, rheumatoidarthritis or neurogenic arthritis, generalised soft tissue rheumatism(fibromyalgia), neurogenic inflammation of the oral mucosa, inflammatorylung diseases, allergic rhinitis, asthma and COPD, diseases accompaniedby excessive vasodilatation and resultant reduced blood supply to thetissues, e.g. in particular shock and sepsis, chronic pain, such as e.g.diabetic neuropathies, neuropathies induced by chemotherapy, HIV-inducedneuropathies, post-herpetic neuropathies, neuropathies induced by tissuetrauma, trigeminal neuralgias, temporomandibular dysfunctions, CRPS,back pain and visceral diseases such as for example IBS (irritable bowelsyndrome) or inflammatory bowel syndrome, for relieving pain in generalor for the preventive or acute therapeutic treatment of the symptoms ofhot flushes caused by vasodilatation and increased blood flow inmenopausal oestrogen-deficient women and hormone-treated patients withprostate carcinoma and castrated men, which comprises the administrationto a person suffering from the same a therapeutic amount of a compoundaccording to claim 1, 2, 3, 4, 5 or 6 or a physiologically acceptablesalt thereof together.